Diet Composition and Feeding Frequency for Jumbo Shiner Production
Nathan Stone, Alex Kachowski, Rebecca Lochmann and Greg O’Neil

A pond study evaluated the effect of diet composition and feeding frequency on the growth and production of golden shiners. Juvenile golden shiners were stocked into 12, 0.04-ha earthen ponds at a rate of 74,100/ha.  Fish were fed either once or twice daily with one of two diets: control diet with fish meal and an experimental diet without fish meal, with the intent of matching the performance of fish fed the control diet but at a lower cost. At harvest, there was no significant difference in production or average weight of fish due to diet or feeding frequency. 

Effect of Dietary Vitamin C Concentration on Growth and Health of Largemouth Bass Micropterus salmoides.
Rebecca Lochmann, Todd Sink and Harold Phillips

The objective was to determine the minimum concentration of vitamin C (ascorbic acid; AA) required in a practical diet to optimize growth and prevent specific signs of AA deficiency in juvenile largemouth bass.17-gram LMB were fed practical diets with different concentrations of vitamin C (0, 25, 50, 100, 250, or 500 mg AA^. kg^-1 diet). Total mass per tank was measured once every two weeks.  No external signs of AA deficiency were observed but growth rates were low, probably due to chronic infestation with leeches. Fish were treated twice during the study with Dylox, but the leeches were never entirely eradicated. Health assays are in progress. Preliminary results indicate that the alternative complement activity of all fish was increased 3-fold compared to fish not infected with leeches. It is probable that any vitamin-C induced increases in complement activity will be obscured by increases due to leech infestation alone. Lysozyme concentrations were also elevated by leech infestation. Fish carcasses will be analyzed for visceral and whole body concentrations of ascorbic acid, protein, lipid, dry matter and ash. Although we cannot determine the dietary requirement for vitamin C due to the compromised performance of the fish, we are trying to determine whether the fishes fed diets with higher concentrations of vitamin C were protected in any way from negative health effects of the leeches. This trial will be treated as a case study – not a nutrient requirement study, and any benefits cannot be determined until all analyses are complete. 

An Enzyme-linked Immunosorbent Assay for Sampling Blood Plasma Insulin Concentrations in Largemouth Bass, Micropterus salmoides.
Rebecca Lochmann, Todd Sink and Harold Phillips

The objective was to validate a non-radioisotopic enzyme-linked immunosorbent assay (ELISA) for detection of plasma insulin concentrations in largemouth bass, Micropterus salmoides. Wild LMB were subjected to oral glucose stimulation or fasted for 2 d. Farm-raised, pellet-trained bass were fed a commercial diet containing approximately 25% soluble carbohydrates. All three groups of fish were then bled and plasma was used for validation of the assay. A solid phase two-site enzyme immunoassay in which two monoclonal antibodies are bound to separate antigens on the insulin molecule was conducted. The results indicate that an ELISA insulin kit could be used for experimental detection of blood plasma insulin concentrations in LMB. The linearity, precision, and reproducibility of the data were acceptable. However, the accuracy is unproven as no “gold standard” for LMB insulin exists. This ELISA method would be most useful for the detection of insulin, general increases or decreases in insulin (insulin response), and general comparative trends in insulin concentration in LMB and possibly other fish. This method will be useful in development of commercial diets for largemouth bass to determine appropriate carbohydrate loads that support growth and health while remaining cost effective.

Insulin Response of Largemouth Bass to Glucose, Amino Acid, and Diet Stimulation.
Rebecca Lochmann, Todd Sink and Harold Phillips

The objective was to characterize the insulin response of largemouth bass to diet, glucose, and amino acids. Pellet-trained largemouth bass were either orally stimulated with glucose, lysine, arginine, methionine, or fed a commercial pelleted diet, or a “natural” diet of golden shiners and bled over time for determination of insulin response. Insulin response was correlated with glucose response, as insulin response lagged slightly behind increases in blood glucose concentration. Greater increases in blood glucose were accompanied by sharper, quicker insulin responses. Glucose (4.8 ± 0.3 ng/mL insulin) and arginine (5.4 ± 0.4 ng/mL insulin) stimulation evoked the greatest mean insulin responses at 60 min post-administration, while lysine (2.1 ± 0.3 ng/mL insulin) also produced elevated insulin responses. The pelleted diet stimulated an intermediate response, while the “natural” diet of shiners produced the lowest insulin response. Due to confounding effects and interference from arginine and lysine in the diet, insulin response is not suitable for determining appropriate carbohydrate loads in largemouth bass diets. At this time, histological data and liver glycogen concentration should be used to determine appropriate carbohydrate loads in commercial diets. This technique may be valuable in assessing optimal carbohydrate loads in commercial largemouth bass diets. 

Validation, Use, and Disadvantages of Enzyme-linked Immunosorbent Assay Kits for Detection of Cortisol in Channel Catfish, Largemouth Bass, Red Pacu, and Golden Shiners.
Rebecca Lochmann, Todd Sink and Harold Phillips

The objective was to validate procedures for multiple commercial cortisol ELISA kits and compare the results to the RIA method. The assays were tested for four species: 1) channel catfish Ictalurus punctatus; 2) largemouth bass Micropterus salmoides; 3) red pacu Piaractus brachypomus; and 4) golden shiners Notemigonus crysoleucas. We evaluated the ELISA methods against RIA, and determined that at least one kit (Assay Designs, Inc. #900-071) is suitable (accuracy: mean recovery of spiked samples, 102.8%; reproducibility: inter-assay coefficient of variation < 10.5% for all species; precision: intra-assay coefficient of variation < 16.8% for all species; linearity: R² > 0.96 for all species) for measurement of cortisol response in fish and comparative determination of stress. All of the ELISA assay results varied more than 10% from the cortisol concentrations detected by the RIA. The high variability of kit results indicate that commercial ELISA kits could be utilized for qualitative determination of cortisol in fish, but should be fully validated in each laboratory for each species before an ELISA kit is used for research. Researchers will have effective means to measure the stress response during experiments with species important to Arkansas and other locations. 

Effect of Practical Carbohydrate Sources from Native Peruvian Feedstuffs on Growth and Health of Piaractus brachypomus and Colossoma macropomum.
Rebecca Lochmann, Todd Sink and Harold Phillips

The objective was to determine performance and health of Characid fishes fed diets containing carbohydrates from different Peruvian feedstuffs.We conducted two separate feeding trials to determine the effects of a control diet (containing wheat) versus diets with one of three native Amazonian plant feedstuffs on the performance of C. macropomum (Trial 1) and P. brachypomus (Trial 2). The native feedstuffs were pijuayo (Bactris gasipaes), plátano (Musa paradisiaca), and yucca (Manihot sculenta), which are all widespread in the Amazon region.  Diets were formulated to contain similar amounts of total protein and energy and differed only in the test feedstuff (wheat, pijuayo, platano or yucca) being evaluated primarily as an energy source. Relative to wheat bran all of the feedstuffs tested were effective energy sources for C. macropomum and P. brachypomus and increased use of pijuayo, plátano, and yucca  may reduce diet cost and enhance sustainability of Characid culture in Amazonia . Similar feed ingredients can be used in other locations where pacu are raised as ornamental fish. Results may be applicable to other warmwater omnivorous fishes that are commercially important in Arkansas .