Role of dietary ingredients on fish performance and genes associated with appetite, growth, metabolism, and intestinal inflammation in channel catfish (Ictalurus punctatus)

In commercial channel catfish production, feed costs are the largest variable expense. In recent years, the use of fishmeal in catfish feeds has raised sustainability and cost concerns, and as such, a notable amount of research has been dedicated to the exploration of alternatively-sourced ingredients for use in channel catfish diets. The present research evaluated the effects of alternatively-sourced dietary ingredients on production indices, gut health, and genes involved in the regulation of appetite, growth, metabolism, and intestinal inflammation. In Objective 1, 45 sets of oligo primers were designed based on sequences in the Genbank database. PCR products from primers designed using teleost sequences outside of the genus Ictaluridae were cloned, transformed, and sequenced to verify gene identity. The entire set of designed primers was then validated for use in real-time PCR and used to validate a high-throughput, microfluidic, dynamic, qPCR array for measuring gene expression in the previously mentioned pathways. Thresholds for efficiency were set at R2 ≥ 0.9 and 0.65 for quality, using a baseline correction and auto detectors for the Ct threshold method. Of the 45 genes analyzed using a Fluidigm qPCR System, eigth genes failed due to early or late amplification, low PCR efficiency, or a low quality score. Of the potential reference genes evaluated, α-tubulin had the highest PCR efficiency (0.9989), a high quality score (0.92±0.03) acceptable Ct values. Based these results, 40 genes from four tissues (hypothalamus/pituitary, liver, stomach, proximal intestine) were evaluated in Objective 2 in response to three alternative dietary treatments, relative to a control diet, using the Fluidigm qPCR System. Changes in dietary formulations were primarily based on alternative protein feedstuffs, with the following feedstuffs as the distinguishing protein sources: (A) fish meal (control), (B) porcine meat and bone meal, (C) soybean meal, and (D) corn germ meal/cottonseed meal. All diets were isonitrogenous and isolipidic. Dietary treatments were fed to three tanks of 20 channel catfish each for a duration of 12 weeks. Differences in feed conversion ratio (FCR), specific growth rate, feed intake, body condition, weight gain, proximal intestine histology, and quantitative gene expression were analyzed. FCR was significantly (P\u3c0.05) increased in Diets 2 – 4 relative to Diet 1; however, other production indices were unaffected by treatment. Dietary treatment also had no effect on intestinal histology (P\u3c0.05). Diets 2, 3, and 4 caused diet specific differences (P\u3c0.05) in the expression of neuropeptide Y, peptide YY, α-amylase, insulin receptor-a, G6PD, GR1, and GR2, relative to the control (Diet 1). These changes likely relate to differences in diet-mediated regulation of appetite and glucose metabolism, and perhaps the modulation of gut passage rate by PYY. By evaluating the molecular regulation of these pathways, effects not detectable in short term feeding trials may be elucidated which effect fish well-being and may explain subtle differences in performance, such FCR, as observed in the present study. This research supports the use of targeted gene panels in aquaculture nutrition, particularly in the development of sustainable catfish feeds