Functional feeds marginally alter immune expression and microbiota of Atlantic salmon (Salmo salar) gut, gill, and skin mucosa though evidence of tissue-specific signatures and host-microbe coadaptation remain.

BACKGROUND: Mucosal surfaces of fish provide cardinal defense against environmental pathogens and toxins, yet these external mucosae are also responsible for maintaining and regulating beneficial microbiota. To better our understanding of interactions between host, diet, and microbiota in finfish and how those interactions may vary across mucosal tissue, we used an integrative approach to characterize and compare immune biomarkers and microbiota across three mucosal tissues (skin, gill, and gut) in Atlantic salmon receiving a control diet or diets supplemented with mannan-oligosaccharides, coconut oil, or both. Dietary impacts on mucosal immunity were further evaluated by experimental ectoparasitic sea lice (Lepeophtheirus salmonis) challenge. RESULTS: Fish grew to a final size of 646.5 g ± 35.8 during the 12-week trial, with no dietary effects on growth or sea lice resistance. Bacterial richness differed among the three tissues with the highest richness detected in the gill, followed by skin, then gut, although dietary effects on richness were only detected within skin and gill. Shannon diversity was reduced in the gut compared to skin and gill but was not influenced by diet. Microbiota communities clustered separately by tissue, with dietary impacts on phylogenetic composition only detected in the skin, although skin and gill communities showed greater overlap compared to the gut according to overall composition, differential abundance, and covariance networks. Inferred metagenomic functions revealed preliminary evidence for tissue-specific host-microbiota coadaptation, as putative microbiota functions showed ties to the physiology of each tissue. Immune gene expression profiles displayed tissue-specific signatures, yet dietary effects were also detected within each tissue and peripheral blood leukocytes. Procrustes analysis comparing sample-matched multivariate variation in microbiota composition to that of immune expression profiles indicated a highly significant correlation between datasets. CONCLUSIONS: Diets supplemented with functional ingredients, namely mannan-oligosaccharide, coconut oil, or a both, resulted in no difference in Atlantic salmon growth or resistance to sea lice infection. However, at the molecular level, functional ingredients caused physiologically relevant changes to mucosal microbiota and host immune expression. Putative tissue-specific metagenomic functions and the high correlation between expression profiles and microbiota composition suggest host and microbiota are interdependent and coadapted in a tissue-specific manner.

Insect (black soldier fly larvae) oil as a potential substitute for fish or soy oil in the fish meal-based diet of juvenile rainbow trout (Oncorhynchus mykiss).

Alternative sources of fish oil (FO) are one of the major problems in aquaculture; therefore, the goal of the present study was to examine insect (black soldier fly larvae) oil (BSLO) as a potential replacer of fish/soy oil in juvenile rainbow trout (initial average weight of 32 ± 0.15 g) feed. Four diets were formulated wherein FO (control diet) was completely replaced with either soybean oil (SO) or BSLO, and an additional BSLO-based diet supplemented with 1.5% bile acid (BSLO + BA) were fed to the fish for 10 weeks. Growth performance of the BSLO fed group was similar (P > 0.05) to that of the FO and SO fed groups, however, the fish fed BSLO + BA diet registered the lowest growth (P < 0.05). Oil sources did not (P > 0.05) affect the major nutrient content of whole-body, however, the fatty acid composition of the muscle and liver was influenced (P < 0.05), with the highest 14:0, 16:0, and total saturated fatty acid detected in BSLO or BSLO + BA fed trout compared to the others (P < 0.001). No significant differences were observed in eicosapentaenoic acid + docosahexaenoic acid (EPA + DHA) or total n-3 polyunsaturated fatty acid (PUFA) content in muscle among the groups, whereas, the highest EPA:DHA and n-3:n-6 ratios were detected in the FO group. Gene expression for fatty acid binding protein (fabp), fatty acid synthase (fas), and Δ5 desaturase in the liver was lower in FO (P < 0.05), while BSLO + BA registered the highest Δ6 expression (P = 0.006). Supplementation of BA in the BSLO diet increased superoxide dismutase (SOD) and catalase (CAT) activities compared to the other groups (P < 0.05). In conclusion, BSLO could serve as a substitute for FO and SO in rainbow trout diet without negatively impacting growth performance, whole-body composition and nutrient retention, and modulate the expression of fatty acid metabolism-related genes in rainbow trout.

Digesta and Plasma Metabolomics of Rainbow Trout Strains with Varied Tolerance of Plant-Based Diets Highlights Potential for Non-Lethal Assessments of Enteritis Development.

The replacement of fishmeal in aquafeeds is essential to the sustainability of aquaculture. Besides the procurement of alternative protein sources, fish can also be selected for better performance on plant-based alternative diets. Rainbow trout (Oncorhynchus mykiss) is one such species in which the strain ARS-Sel has been selected for higher growth and enhanced utilization when fed soy-based diets. The aim of this study was to compare fish growth and plasma and digesta metabolomes between ARS-Sel and two commercial strains (CS-1 and CS-2), when fed plant-protein (PM) and fishmeal-based (FM) diets, and to correlate them with the onset of enteritis. An NMR-metabolomics approach was taken to assess plasma and digesta metabolite profiles. Diet and strain showed significant effects on fish growth, with the ARS-Sel fish receiving the PM diet reaching the highest final weight at sampling. Multivariate analysis revealed differences between plasma and digesta metabolite profiles of ARS-Sel and CS (CS-1 considered together with CS-2) PM-fed groups in the early stages of enteritis development, which was confirmed by intestinal histology. As reported in previous studies, the ARS-Sel strain performed better than the commercial strains when fed the PM diet. Our findings also suggest that metabolomic profiles of plasma and digesta, samples of which can be obtained through non-lethal methods, offer valuable insight in monitoring the occurrence of enteritis in carnivorous aquaculture species due to plant-based diets.

Chronic exposure to environmental cadmium affects growth and survival, cellular stress, and glucose metabolism in juvenile channel catfish (Ictalurus punctatus).

Anthropogenic activities have led to the enrichment of cadmium in freshwater systems where it is a contaminant of concern for fisheries and aquaculture as it has no known biological function and is toxic at trace concentrations. Yet, knowledge gaps remain regarding effects of chronic exposure to environmentally relevant concentrations on freshwater fish. Thus, the objectives of the current study were to assess chronic impacts of cadmium on channel catfish (Ictalurus punctatus) including how tissue-specific bioaccumulation patterns relate to functions of those tissues over time. We focused on liver and kidneys, and expression of genes related to cellular stress, glucose metabolism, and steroidogenesis. Catfish were exposed to concentrations of 0.5 (control), 2 (low), and 6 (high) µg L-1 Cd from fertilization to six months. Cadmium exposure negatively impacted channel catfish growth and was linked to bioaccumulation of tissue Cd, which followed a dose-related response, where concentrations in trunk kidney > liver = head kidney >> muscle. Differences in tissue Ca, Cu, Fe, and Zn concentrations were also observed between treatments. Following 3 months of exposure, expression of metallothionein (MT) and heat shock proteins (HSP) 70 & 90 increased relative to controls; however, no differences were detected at 6 months, suggesting compensation. Conversely, there were no differences in expression patterns for key genes in steroidogenesis, steroidogenic factor 1 (SF1), steroidogenic acute regulatory protein (StAR), and cytochrome P450scc (P450), which supports the observation that Cd did not affect the secondary stress response, evaluated via plasma cortisol and glucose concentrations following a low water stress event. As a function of length and weight, the high Cd treatment yielded fish that were significantly smaller than controls. In addition to the cellular responses in MT and HSPs noted, reduced growth in the high Cd treatment was likely due, at least in part, to elevated energetic demands. This is supported by observations of the upregulation of genes necessary for glucose metabolism. Hexokinase (HK), glucose-6-phosphatase (G6P), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were significantly elevated in the high treatment relative to controls at 3 months of exposure. Over the study period, exposure also reduced survival of channel catfish from 3 to 6 months. Reduced fitness, as a consequence of cadmium exposure, could be visible at the population level through altered life histories and growth patterns.

Insect (black soldier fly, Hermetia illucens) meal supplementation prevents the soybean meal-induced intestinal enteritis in rainbow trout and health benefits of using insect oil.

Black solider fly larvae (BSFL) and their oils (BSFLO) are receiving increasing attention as sustainable ingredients in fish feeds, but mostly as replacements to marine sources. There were two aims to this study; in exp. 1, soybean meal (SBM)-based diets were formulated to contain BSFL as supplements at 0 (SBM), 8 (SBM + BSFLlow) or 16% (SBM + BSFLhigh) with a control diet being fishmeal-based (FM). In exp. 2, diets included only fish oil (FO), soybean oil (SBO), BSFLO or BSFLO + bile acid (BA), and all lipid sources were added at 16%. Both experiments were run at the same time and fed to rainbow trout (32 g) with each treatment being triplicated. After 10 weeks the fish were sampled for liver and distal intestine histology, expression of genes responsible for inflammation in the intestine and kidneys, and serum peroxidase and lysozyme activities. In exp. 1, supplementations of BSFL effectively prevented SBM-induced intestinal enteritis, down-regulated intestinal prostaglandin and interferon regulatory factor 1 (IRF-1), while the SBM + BSFLhigh diet significantly increased serum lysozyme activity. In exp. 2, BSFLO caused no histomorphological change to the liver or intestine, but kidney interluekin-8, tumor necrosis factor and IRF-1 were significantly upregulated along with significantly higher serum peroxidase activity. The inclusion of BA in the BSFLO diets significantly upregulated intestinal prostaglandin gene expression. Overall, BSFL supplementations of 8 or 16% prevented SBM-induced intestinal enteritis based on histological observations, which was supported by a down-regulation in pro-inflammatory genes and enhanced innate immunity. Meanwhile, the use of BSFLO showed some immunological benefits. Therefore, these sustainable resources are recommended in the diets of rainbow trout, especially when using elevated levels of plant-based proteins.

Ontogeny of the cortisol stress response and glucocorticoid receptor expression during early development in channel catfish, Ictalurus punctatus.

Cortisol is a glucocorticoid hormone which is an endocrine signaling molecule in all vertebrates and acts through intracellular glucocorticoid receptors (GRs). Cortisol affects many biological functions including immunity, stress, growth, and reproduction. The objective of this study was to investigate the ontogeny of the cortisol and GR stress response in channel catfish (Ictalurus punctatus) at several early life stages. To accomplish this, resting and stress-induced levels of tissue cortisol and the two catfish GRs (GR-1 and GR-2) expression were measured. Resting cortisol levels in newly fertilized eggs averaged 2.4 ±â€¯0.2 ng/egg and decreased to 0.4 ±â€¯0.01 ng/egg by day 5. Cortisol levels in newly fertilized eggs subjected to an acute stress (lowered dissolved oxygen from 6.5 mg/L to 1.8 mg/L) averaged 2.3 ±â€¯0.1 ng/egg and decreased to 0.3 ±â€¯0.03 ng/egg by day 5. At hatching, resting cortisol levels were 24 ±â€¯1.0 ng/0.1 g tissue while levels increased to 83 ±â€¯2.0 ng/0.1 g tissue in fry subjected to an acute stress (P < .05). Four days post-hatch, resting cortisol levels were 83 ±â€¯1.0 ng/0.1 g tissue while levels increased to 149 ±â€¯4.0 ng/0.1 g tissue in fry subjected to an acute stress (P < .01). There was no significant difference between GR-1 and GR-2 mRNA in stressed and unstressed newly hatched fry. Four days post-hatch, GR-1 mRNA increased 3-fold while GR-2 mRNA increased 2-fold in fry that were subjected to low dissolved oxygen conditions (P < .05). These results indicate that cortisol biosynthesis, integration and maturation of the hypothalamic-pituitary-interrenal (HPI) axis can be observed in channel catfish at hatching. The upregulation of GR-1 and -2 mRNA in stressed fry supports roles for both transcripts in integrating the channel catfish stress response.

Exposure to environmentally relevant cadmium concentrations negatively impacts early life stages of channel catfish (Ictalurus punctatus).

Cadmium is a persistent contaminant of surface waters. The effects of cadmium on early life stages of fish are not well understood, although they are often disproportionately affected by contaminants. The objectives of this study were to examine effects of chronic exposure to environmentally relevant concentrations on growth, development, cellular stress, and glucose metabolism of channel catfish, Ictalurus punctatus. Eggs were wet-fertilized in treatment water at concentrations of 0.4 (control), 2.2 (low), or 8.5 (high) µg L-1 and monitored through swim-up, black fry stage. Eggs and fry accumulated cadmium dose-dependently. Fertilization rates were unaffected, yet hatch rate was significantly reduced in the high treatment. Survival to black fry and overall size and condition factor were not affected; however, differences in yolk sac size, and presumably energetics of yolk fry, was detected. Physiological pathways were also affected, demonstrated by altered gene expression, most notably in genes related to carbohydrate metabolism. Elevated expression of HK and G6PD, rather than G6P and GADPH, suggests glucose may be shunted towards the pentose-phosphate pathway. Overall, observations indicate cadmium negatively affects development in early life stages of channel catfish, which could lead to shifts in population structure and life history patterns in exposed populations of wild fish.

Rapid acclimation of the cortisol stress response in adult turquoise killifish Nothobranchius furzeri.

The turquoise killifish Nothobranchius furzeri is an increasingly popular model species for comparative vertebrate research, and the basic physiology including responses to stressful stimuli are of primary interest. We exposed adult killifish to a single or repeated periods of acute confinement followed by analysis of tissue cortisol and plasma cortisol concentrations. Individuals were also sampled for messenger RNA (mRNA) expression of corticotropin-releasing hormone (CRH), mineralocorticoid receptor (MR), and glucocorticoid receptor (GR) in the brain to examine the effects of repeated stress events on constitutive expression of these important stress axis components. Following a single 30-minute confinement stress, male plasma cortisol significantly differed from baseline (p = 0.04). Both male and female whole-body cortisol were significantly increased (p = 0.004 and p = 0.04, respectively) at 15 and 30 minutes poststress. Despite obvious dimorphic behavior and morphology, cortisol concentrations did not differ between the sexes. Exposure to daily repeated confinement for one week altered the cortisol response in both sexes. Time 0, 15, and 60 minutes poststress cortisol concentrations were depressed in repeatedly stressed males (p ≤ 0.05), and times 0, 30 and 120 minutes poststress cortisol concentrations were depressed in repeatedly stressed females (p ≤ 0.05). Constitutive expression of CRH, MR, and GR mRNA in the brain following one week of repeated stress events did not differ among treatments or sexes. This study introduces the first description of hypothalamic-pituitary-interrenal axis activity in this important model species. Reduced cortisol production in repeatedly stressed adult killifish suggests acclimation to repeated stressors. Furthermore, acclimation was rapid, and plasma cortisol concentrations altered significantly in as little as one week.

Characterization of a third ghrelin receptor, GHS-R3a, in channel catfish reveals novel expression patterns and a high affinity for homologous ligand.

A novel third channel catfish growth hormone secretagogue (ghrelin) receptor, GHS-R3a, gene was characterized. Identification and analysis of the genomic organization of channel catfish GHS-R3a revealed differences in exon/intron structure relative to the previously published GHS-R1a and GHS-R2a sequences. Amino acid sequence alignment of catfish GHS-R3a with -R1a and -R2a revealed 48 and 52% sequence identity, respectively. Phylogenetic analysis predicted a new clade of GHS-R3a receptors found only in fish, with representation in the teleost infradivisions Osteoglossomorpha, Clupeomorpha, and Euteleostei. In functional analyses, homologous catfish ghrelin increased intracellular Ca2+ concentration in human embryonic kidney (HEK) 293 cells stably expressing catfish GHS-R3a. On the contrary, intracellular Ca2+ concentration was unaffected by treatment with the synthetic growth hormone secretagogues GHRP-6 and hexarelin. Realtime PCR results indicated high expression of GHS-R3a in the brain and gonads, demonstrating tissue specificity among the catfish GHS-Rs. The effects of fasting and refeeding on all three ghrelin receptors were evaluated in catfish brain, pituitary, stomach, and Brockmann bodies. Most notably, GHS-R3a was the only receptor observed to significantly increase (2.9-6.3-fold) in brain, pituitary, and stomach within 4 days of fasting (P < .05). Stomach GHS-R1a also increased (P < .05) after 4 days; however, GHS-R2a was only elevated in brain and pituitary after refeeding for 1 week. Expression of all three ghrelin receptors were elevated (P < .05) in the Brockmann bodies after 2 weeks of fasting and returned to prefasting levels following refeeding. Together with the previously published characterization of GHS-R1a and -R2a, these results establish three ghrelin receptors, each altered by energy state, in channel catfish and add to the growing body of information on GHS-R evolution and function.

Comparison of Channel Catfish and Blue Catfish Gut Microbiota Assemblages Shows Minimal Effects of Host Genetics on Microbial Structure and Inferred Function.

The microbiota of teleost fish has gained a great deal of research attention within the past decade, with experiments suggesting that both host-genetics and environment are strong ecological forces shaping the bacterial assemblages of fish microbiomes. Despite representing great commercial and scientific importance, the catfish within the family Ictaluridae, specifically the blue and channel catfish, have received very little research attention directed toward their gut-associated microbiota using 16S rRNA gene sequencing. Within this study we utilize multiple genetically distinct strains of blue and channel catfish, verified via microsatellite genotyping, to further quantify the role of host-genetics in shaping the bacterial communities in the fish gut, while maintaining environmental and husbandry parameters constant. Comparisons of the gut microbiota among the two catfish species showed no differences in bacterial species richness (observed and Chao1) or overall composition (weighted and unweighted UniFrac) and UniFrac distances showed no correlation with host genetic distances (Rst) according to Mantel tests. The microbiota of environmental samples (diet and water) were found to be significantly more diverse than that of the catfish gut associated samples, suggesting that factors within the host were further regulating the bacterial communities, despite the lack of a clear connection between microbiota composition and host genotype. The catfish gut communities were dominated by the phyla Fusobacteria, Proteobacteria, and Firmicutes; however, differential abundance analysis between the two catfish species using analysis of composition of microbiomes detected two differential genera, Cetobacterium and Clostridium XI. The metagenomic pathway features inferred from our dataset suggests the catfish gut bacterial communities possess pathways beneficial to their host such as those involved in nutrient metabolism and antimicrobial biosynthesis, while also containing pathways involved in virulence factors of pathogens. Testing of the inferred KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways by DESeq2 revealed minor difference in microbiota function, with only two metagenomic pathways detected as differentially abundant between the two catfish species. As the first study to characterize the gut microbiota of blue catfish, our study results have direct implications on future ictalurid catfish research. Additionally, our insight into the intrinsic factors driving microbiota structure has basic implications for the future study of fish gut microbiota.

Characterization of two channel catfish, Ictalurus punctatus, glucocorticoid receptors and expression following an acute stressor.

Two channel catfish glucocorticoid receptor genes, ipGR1 (NR3C1_1) and ipGR2 (NR3C1_2) were partially characterized. Identification and analysis of the genomic organization of two channel catfish glucocorticoid (GC) receptors (GRs) revealed differences in the lengths of exons 1 and 2 and the addition of an extra 27-bp exon inserted after exon 2 in the GR1 gene, yielding a 9-aa insert in the receptor protein. Sequence of the 9-aa insert in ipGR1 (WRARQNTHG) is unique compared to other teleost fish GRs. Amino acid sequence alignment of the two channel catfish GRs, revealed 55% sequence identity between them, with a high degree of sequence conservation (82%) in the DNA binding and ligand binding domains. Real-time PCR indicated that ipGR1 and ipGR2 were expressed in all tissues evaluated. Channel catfish GR1 was predominantly expressed in the gills, nearly 25-fold higher than in the liver. GR1 expression was higher than GR2 expression in gills, intestine, head kidney and heart (P<0.05). Channel catfish hepatic GR1 mRNA expression was significantly (P<0.05) increased from pre-stress expression 30min following removal of the acute stressor. After 30min of stress and during the 2h recovery period, ipGR1 mRNA expression was higher relative to ipGR2 expression. Hepatic ipGR2 expression was not affected (P>0.05) by the acute stress event. The present study adds to the growing body of information on GR evolution and function and further demonstrates the unique regulation of the GC/GR system in teleost fish.

Ontogenetic Characterization of the Intestinal Microbiota of Channel Catfish through 16S rRNA Gene Sequencing Reveals Insights on Temporal Shifts and the Influence of Environmental Microbes.

Aquaculture recently overtook capture fisheries as the largest producer of food fish, but to continue increasing fish production the industry is in search of better methods of improving fish health and growth. Pre- and probiotic supplementation has gained attention as a means of solving these issues, however, for such approaches to be successful, we must first gain a more holistic understanding of the factors influencing the microbial communities present in the intestines of fish. In this study, we characterize the bacterial communities associated with the digestive tract of a highly valuable U.S. aquaculture species, channel catfish Ictalurus punctatus, over the first 193 days of life to evaluate temporal changes that may occur throughout ontogenetic development of the host. Intestinal microbiota were surveyed with high-throughput DNA sequencing of 16S rRNA V4 gene amplicons derived from fish at 3, 65, 125, and 193 days post hatch (dph), while also characterizing the environmental microbes derived from the water supply and the administered diets. Microbial communities inhabiting the intestines of catfish early in life were dynamic, with significant shifts occurring up to 125 dph when the microbiota somewhat stabilized, as shifts were less apparent between 125 to 193 dph. Bacterial phyla present in the gut of catfish throughout ontogeny include Bacteroidetes, Firmicutes, Fusobacteria, and Proteobacteria; with the species Cetobacterium somerae and Plesiomonas shigelloides showing the highest abundance in the catfish microbiota after 3 dph. Comparisons of the gut microbiota to the environmental microbes reveals that the fish gut is maintained as a niche habitat, separate from the overall microbial communities present in diets and water-supply. Although, there is also evidence that the environmental microbiota serves as an inoculum to the fish gut. Our results have implications for future research related to channel catfish biology and culture, and increase our understanding of ontogenetic effects on the microbiota of teleost fish.

Functional characterization of insulin-like growth factors in an ancestral fish species, the Shovelnose sturgeon Scaphirhynchus platorhynchus.

Observations from the present study provide the first characterization of the GH-IGF axis in Shovelnose sturgeon Scaphirhynchus platorhynchus, an ancestral fish species. An initial characterization of steady-state IGF-I and IGF-II gene expression in multiple tissues was conducted using real-time RT-qPCR. Overall, the tissues had significantly different profiles of IGF-I gene expression, with the highest IGF-I expression observed in the liver. The highest IGF-II gene expression was also observed in the liver, with minimal or no detection in muscle. A comparison between IGF-I and IGF-II expression within individual tissues revealed higher levels of IGF-II than IGF-I mRNA in the spleen, stomach and trunk kidney, and higher levels of relative IGF-I mRNA expression in the intestine and muscle. The GH-IGF axis was further elucidated by observing the effects of exogenous GH on IGF-I and IGF-II expression in liver and muscle tissue. The results revealed a significant dose-dependent response of both hepatic IGF-I and IGF-II, and muscle IGF-I mRNA expression following rbGH administration. At the highest rbGH concentration (240µg/g BW), IGF-I mRNA levels in liver and muscle peaked significantly at 48h, indicating both hepatic and muscle IGF-I expression to be stimulated by GH. Hepatic IGF-II expression was also stimulated 48h following rbGH administration. Expression of IGF-II mRNA was not inducible in the muscle. Few studies have evaluated the effects of exogenous GH on IGF expression in ancestral vertebrate species, and as such, this research provides valuable insight into the evolution of the somatotropic axis in vertebrates.

Elucidating the roles of gut neuropeptides on channel catfish feed intake, glycemia, and hypothalamic NPY and POMC expression.

Both intrinsic and extrinsic factors modulate food intake and glycemia in vertebrates, in part through interactions with hypothalamic neuropeptide Y (NPY) and proopiomelanocortin (POMC) neurons. The objective of this project was to elucidate the effects of ghrelin (GHRL), gastrin-releasing peptide (GRP), cholecystokinin (CCK), glucagon-like peptide (GLP), pancreatic polypeptide (PP), and peptide YY (PYY) on appetite, glycemia, and hypothalamic expression of NPY and POMC in channel catfish. Catfish were injected intraperitoneally with a single peptide at concentrations of either 0 (control), 50, 100, or 200 ng/g body weight (BW), respectively. Fish were allowed to recover for 30 min, and then fed to satiation over 1 h. Feed intake was determined 1h post-feeding. Catfish injected with GHRL at 50 and 100 ng/g BW and GRP at 200 ng/g BW consumed significantly (P<0.05) less feed compared to controls. A tendency (P<0.1) to suppress feed intake was also observed in the 200 ng/g BW GHRL and PP treatments. PYY, CCK, and GLP had no effects on feed intake. Glycemia was not affected by GHRL, GRP, PP, and PYY treatments, but was suppressed by CCK. A tendency toward lower plasma glucose concentrations was observed in fish administered GLP at 50 ng/g BW. Hypothalamic NPY expression was highly variable and not significantly affected by treatment. POMC expression was also variable, but tended to be reduced by the highest concentration of CCK. These results provide new insight into the roles and regulation of gut neuropeptides in catfish appetite and glycemia.

Exogenous recombinant bovine growth hormone stimulates growth and hepatic IGF expression in shovelnose sturgeon Scaphirhynchus platorhynchus.

Sturgeon are a unique fish for physiological research as they are long-lived, slow-growing, and late-maturing. Furthermore, sturgeon growth hormones appear to share greater structural and molecular similarity with mammalian somatotropins than teleostean somatotropins. In this study, changes in insulin-like growth factor (IGF)-I and IGF-II mRNA expression and corresponding whole-body growth and composition following 6 weeks of bi-weekly recombinant bovine growth hormone (rbGH) administration in shovelnose sturgeon Scaphirhynchus platorhynchus were evaluated. Fish were injected intraperitoneally with 240 µg rbGH/g body weight or a sesame oil sham. Hepatic IGF-I and IGF-II mRNA abundance was significantly higher (P≤0.02) in rbGH-treated fish, as were length (P<0.001) and weight gain (P<0.001). In addition, proximate whole-body analysis demonstrated a significant (P<0.05) increase in protein composition of rbGH-treated fish versus sham-treated fish. There were no significant differences in whole-body moisture, lipid, or ash between the two treatments. These results demonstrate functional roles for GH and IGFs in the promotion of lean growth within this ancient fish species and support the view that the functional effects of GH on hepatic IGF-I expression and somatic growth are conserved from chondostrean to teleostean fishes.

Effects of dietary arginine on endocrine growth factors of channel catfish, Ictalurus punctatus.

Thyroid (TH) and growth (GH) hormones, and insulin-like growth factor I (IGF-I) are anabolic regulators in fish and responsive to nutrient intake. A study was conducted to determine if previously reported growth effects of dietary arginine (ARG) in channel catfish were related to the activation of endocrine axes. In a first experiment, catfish were fed incremental levels of ARG (0.5 - 4% of diet) for 6 weeks and sampled at 2-week intervals. In a second experiment, fasted (48h) fish were fed a single ration of ARG (0.5 or 4% of diet) and sampled at various intervals (0 to 72h postprandial, PP). Experiment 1 did not reveal any influence of ARG on circulating TH, GH, or IGF-I despite the significantly increased growth of fish fed ARG-enriched diets. In experiment 2, feeding the 4% ARG diet significantly increased the amplitude of pulsatile plasma GH levels and also significantly increased IGF-I mRNA in liver and muscle, (at 2h PP) and plasma IGF-I levels (at 6h PP). Although relatively infrequent sampling failed to reveal alterations in TH or GH levels in response to ARG-induced growth activation, PP high frequency sampling unveiled high amplitude pulsatile GH secretions and may be important in activating IGF production in target tissues. Additionally, expressed and secreted IGF-I exhibited discernible patterns which closely correlate with ARG-induced growth effects in catfish.

Pre- and postprandial changes in orexigenic and anorexigenic factors in channel catfish (Ictalurus punctatus).

Ghrelin (GRLN), cocaine and amphetamine regulated transcript (CART), neuropeptide Y (NPY), and cholecystokinin (CCK) are neuropeptides involved in the regulation of appetite and feeding in vertebrates. We examined pre- and postprandial changes in the expression of plasma GHRL and mRNAs encoding GRLN, CART, NPY, and CCK in channel catfish. Fish were entrained to eat at 0900 h for 2 weeks. Fish were then sampled at 0700, 0800, and 0900 h. Remaining fish were either offered feed at 0900 h (Fed) or fasted (Unfed). Fish sampling continued at 0.5, 1, 2, and 4 h post feeding. Feeding increased abundance of whole brain CART mRNA out to 4 h with no effect observed in unfed fish. Whole brain NPY expression peaked at 0.5 h in both treatments. NPY expression then declined in fed fish but remained elevated in unfed fish. No differences in plasma or stomach GRLN expression were observed. Two separate cDNAs for CCK were identified. Brain CCKa and CCKb expression increased after feeding. These results suggest CART, NPY, and CCK play roles in the regulation of channel catfish feeding. Taken together, these results provide new insights into the neural and gastroenteric mechanisms regulating appetite in channel catfish.

Assembly of 500,000 inter-specific catfish expressed sequence tags and large scale gene-associated marker development for whole genome association studies.

BACKGROUND: Through the Community Sequencing Program, a catfish EST sequencing project was carried out through a collaboration between the catfish research community and the Department of Energy's Joint Genome Institute. Prior to this project, only a limited EST resource from catfish was available for the purpose of SNP identification. RESULTS: A total of 438,321 quality ESTs were generated from 8 channel catfish (Ictalurus punctatus) and 4 blue catfish (Ictalurus furcatus) libraries, bringing the number of catfish ESTs to nearly 500,000. Assembly of all catfish ESTs resulted in 45,306 contigs and 66,272 singletons. Over 35% of the unique sequences had significant similarities to known genes, allowing the identification of 14,776 unique genes in catfish. Over 300,000 putative SNPs have been identified, of which approximately 48,000 are high-quality SNPs identified from contigs with at least four sequences and the minor allele presence of at least two sequences in the contig. The EST resource should be valuable for identification of microsatellites, genome annotation, large-scale expression analysis, and comparative genome analysis. CONCLUSIONS: This project generated a large EST resource for catfish that captured the majority of the catfish transcriptome. The parallel analysis of ESTs from two closely related Ictalurid catfishes should also provide powerful means for the evaluation of ancient and recent gene duplications, and for the development of high-density microarrays in catfish. The inter- and intra-specific SNPs identified from all catfish EST dataset assembly will greatly benefit the catfish introgression breeding program and whole genome association studies.

Evaluation of sodium carbonate peroxyhydrate as a potential catfish egg disinfectant.

Two experiments were conducted to evaluate the efficacy of sodium carbonate peroxyhydrate (SCP) in improving the hatching success of channel catfish Ictalurus punctatus when used as a prophylactic chemotherapeutant during egg incubation. In the first experiment, the efficacy of SCP was evaluated in 379-L aluminum incubation troughs similar to those used in commercial hatcheries. Egg masses treated daily with 254 mg of SCP/L of water had significantly higher mean hatching success than untreated controls, and a pathogen-inhibiting effect was also evident (i.e., no gross infection was observed on the treated egg masses). In the second experiment, the hatching success of egg masses treated daily with 254 mg/L was compared with that of egg masses treated daily with hydrogen peroxide (70 mg/L). The effects of both treatments on the pH, dissolved oxygen, and hydrogen peroxide concentrations in the trough were also examined. Both SCP and hydrogen peroxide significantly improved hatching success. Unlike in the treatment with hydrogen peroxide, water pH increased during the treatment with SCP; however, no negative effects on hatching success were observed. The results of this research suggest that SCP acts similarly to hydrogen peroxide in improving channel catfish hatching success and warrants further research to determine whether it could be a practical and effective alternative for managing catfish egg infections in commercial hatcheries.

Sequence, genomic organization and expression of two channel catfish, Ictalurus punctatus, ghrelin receptors.

Two ghrelin receptor (GHS-R) genes were isolated from channel catfish tissue and a bacterial artificial chromosome (BAC) library. The two receptors were characterized by determining tissue distribution, ontogeny of receptor mRNA expression, and effects of exogenous homologous ghrelin administration on target tissue mRNA expression. Analysis of sequence similarities indicated two genes putatively encoding GHS-R1 and GHS-R2, respectively, which have been known to be present in zebrafish. Organization and tissue expression of the GHS-R1 gene was similar to that reported for other species, and likewise yielded two detectable mRNA products as a result of alternative splicing. Expression of both full-length, GHS-R1a, and splice variant, GHS-R1b, mRNA was highest in the pituitary. Gene organization of GHS-R2 was similar to GHS-R1, but no splice variant was identified. Expression of GHS-R2a mRNA was highest in the Brockmann bodies. GHS-R1a mRNA was detected in unfertilized eggs and throughout embryogenesis, whereas GHR-R2a mRNA was not expressed in unfertilized eggs or early developing embryos and was the highest at the time of hatching. Catfish intraperitoneally injected with catfish ghrelin-Gly had greater mRNA expression of GHS-R1a in pituitaries at 2 h and Brockmann bodies at 4 h, and of GHS-R2a in Brockmann bodies at 6 h post injection. Amidated catfish ghrelin (ghrelin-amide) had no observable effect on expression of either pituitary receptor; however, GHS-R1a and GHS-R2a mRNA expression levels were increased 4 h post injection of ghrelin-amide in Brockmann bodies. This is the first characterization of GHS-R2a and suggests regulatory and functional differences between the two catfish receptors.