Description of Concept: We have been able to systematically improve the sustainability and cost effectiveness of practical diets designed for the Florida pompano. Across these studies we have reduced the level of animal proteins from over 30% of the diet to less than 10% while increasing the level of soy products to around 50% of the diet. During this period, we have identified minor changes in gut histology, but no long-term growth trials have been conducted to validate a lack of any adverse effects. Based on previous research we feel that the next step is two-fold. First is to optimize the type of soy used in these formulations. Secondly, we need to conduct longer term growout trials to validate our open soy-based feed formulation against commercial feeds.

Objective 1. Validate our soy-optimized open feed formulation against the commercial feed currently used for this species; Objective 2. Compare the biological value of standard SE Soybean meal to two new genetically improved meals (High protein SBM 58% protein & ultra-low oligosaccharide meal) currently being commercialized by Benson Hill.

The proposed project will result in a series of papers and presentations at trade meetings that will promote the use of soy-optimized feeds. The research is geared to improve the performance of fish on high soy dies. Thus, providing scientific data towards the continued development of soy-based feeds and demonstrate their efficacy to the producer.

Updated March 20, 2022:
The research is seasonal, so we are in the process of planning and working to rebuild systems. Presently, we have evaluated options for obtaining Florida pompano for this year’s work. Unfortunately, the first group of fish we identified and were coordinating pickup were euthanized due to miss communication. A new group of larvae are currently being reared and will be ready by early summer for work. We have initiated the shipping of new soy varieties and have one in stock. We have completed an inventorying our ingredients in preparation for formulating the feeds and making them. In general, we are moving towards the organization of the research over the summer. Hopefully, the delay in fish will not push the project back to far.

Updated June 17, 2022:
The test diets for the growth trial have been formulated and made. The juvenile pompano were received and nursed for around one month. The fish were then size sorted and stocked on 6/14/2022 at a mean weight of 7.8g mean weight. The growth trial will be run for around 8 weeks.

Updated September 26, 2022:
A series of growth trials were conducted to a) compare a high soy low animal meal diet to a commercial feed formulation and b) evaluate the efficacy of new soy-products in diet for the Florida pompano. Diets for these trials were formulated and analyzed, for brevity this data will be presented in the final report.

The growout trials comparing our experimental diet (8% poultry meal, 10% corn protein concentrate, 55% soybean meal) to a commercial feed was conducted in two different recirculation systems. Juvenil fish (23.2g) were reared in our smaller tank (800 L) recirculation system using 6 replicates for each diet. At the conclusion of the 6-week growth final weight (55.2 and 60.4g, P=0.0758), percent weight gain (138.8% and 160.0%; P=0.057) and FCR (2.05, 1.86; P= 0.0153) for fish reared on the experimental feed were generally poor than that of fish reared on the commercial feed, respectively. As similar response was observed in our large tanks system (3,000 L) which utilized three replicate groups of larger fish with an initial mean weight of 38.1g. At the conclusion of the 8 week growth trial final weights (125g and 137g; P-value 0.3318), percent weight gain (226.1% and 269.2 %; P = 0.2836) and FCR ( 2.05 and 1.89; p= 0.5769) were not statistically different albeit fish offered the experimental diet had poorer values. In both cases, performance was poorer, albeit not necessarily significantly (P < 0.05) in fish offered the experimental diet as compared to the commercial feed. This would indicate that although we were close in terms of performance, we still need to optimize diets that have low levels of animal protein and high levels of plant-based proteins.

The second component of the research was to evaluate the use of new soybean sources in our moderate animal protein diet (14 % poultry meal, 6% corn protein concentrate and 50% sovent extracted soybean meal) For this work 4 different soybean types, were used to replace 50 or 100% of the solvent extracted soybean meal in the basal diet. This trial had numerous issues, first we had two electrical outages that caused dissolved oxygen stress to the fish. During the outages, we only lost a few fish, but this probably stressed the fish and predisposing them to disease issues. After the stress the fish experienced a slow mortality, which was not alleviated with therapeutic treatment of the fish and resulted in low survival at the conclusion of the trial. This could have been further complicated by an error in the formulations resulting in taurine not being supplemented to these diets. Based on the analyzed values, there was a low level of taurine in the diets which in the past has not resulted in high mortality but is likely to of added to the stress. At the conclusion of the trial, survival in this trial ranged from 33.7% to 57.5%, well below acceptable levels. Unfortunately, the fish must be reared for these trials so other cohorts of fish were not available nor was there adequate time at the facility to re-run the trial. As an alternative, to no data, we have transferred the fish from our growout trial in Gulf Shores to Auburn where we have large indoor facilities, and we will use a reduced set of diets (100% replacements) and run another trial with larger fish. This trial will be stocked 9/25/2022.

Because of the issues with fish survival, we have requested a no cost extension and we have planned to initiate a fourth growth trial which will be run over the next three months. If the trail is successful, we will then collect blood, liver and distal intestine samples for analysis.

Updated May 19, 2023:
As previously reported, due to system failure in our growout trial for the various soy sources we had poor survival of the fish. Hence, we chose to re-run this trial moving the work to the E. W. Shell Fisheries Laboratory where we have a large RAS system that can be run with artificial sea salt during the winter. We were able to obtain fish from ProAquatix on November 3rd. We acclimated the animal until November 21st and then initiated a growth trial. Because this system has fewer tanks, we had to reduce the number of treatments so the expeller pressed product was not run. This trial included 6 diets including the basal diet and diet for which Bright day (50, 100%) and Hamlet (50, 100%) and Soycomil (100%) were used to replace solvent extracted soybean meal. Other than dropping one soy source the diets we identical to those run previously. After two weeks of the growth trial fish were doing well but then between week two and four they developed signs of Amyloodinium infection. This is a parasite that is typically present but using low doses of copper to kill the dinoflagellate stage and freshwater dips when weighing the fish we typically can keep this under control. However, in this case even high levels of Cu (1 ppm) in the system did not prevent it from reproducing and infecting the fish.

As of December 22, we have less than 50% survival in the fish (due to amyloodinium). We have chosen to move the fish, treat with formalin for the parasite, and re-stock into a different system. That way we can hopefully remove the parasites and be in a system that does not have cysts in it. We have maintained the treatments so it may be possible to rear the remaining fish for histology work. Unfortunately, we have simply run into issues with the growout trials. In January, we will look to see what can be done to salvage as much as possible form this portion of the project.

Updated March 29, 2023:

As previously reported, the original trials were run over the summer of 2022 consisting of two components. The first was run successfully, whereas the second had issues to due electrical failure. Because of issues with the second component of this work a no-cost extension was requested for 6 months and I have put in a second request for another extension to allow completion of the research (see below).

In the first objective, the growout trials comparing our experimental diet (8% poultry meal, 10% corn protein concentrate, 55% soybean meal) to a commercial feed was conducted in two different recirculation systems. Juvenil fish (23.2g) were reared in our smaller tank (800 L) recirculation system using six replicates for each diet. At the conclusion of the 6-week growth final weight (55.2 and 60.4g, P=0.0758), percent weight gain (138.8% and 160.0%; P=0.057) and FCR (2.05, 1.86; P= 0.0153) for fish reared on the experimental feed were poorer but not necessarily significantly than that of fish reared on the commercial feed. A similar response was observed in our large tanks system (3,000 L) which utilized three replicate groups of larger fish with an initial mean weight of 38.1g. At the conclusion of the 8 week growth trial final weights (125g and 137g; P-value 0.3318), percent weight gain (226.1% and 269.2 %; P = 0.2836) and FCR ( 2.05 and 1.89; p= 0.5769) were not statistically different albeit fish offered the experimental diet had poorer values. In both cases, performance was lower, albeit not necessarily significantly (P < 0.05) in fish offered the experimental diet as compared to the commercial feed. This would indicate that although we were very close in terms of performance, we still need to optimize diets that have low levels of animal protein and high levels of plant-based proteins. As the second component of this research was not successfully completed, we chose to take preserved sample from the large tank trial to evaluate histological changes in gut morphology. These samples have been section and we are in the process of evaluating the slides. One of three reviewers have scored the slides and found not significant differences or signs of enteritis. Once, all three reviewers have scored the slides, we will have basic data on whether enteritis occurs in Florida pompano.

The second component of this research was to evaluate various soybean sources. As noted above, the original trial was not successful as we had an electrical failure which stressed the fish resulting in poor survival. A second run was then initiated but we ran into disease issues with amyloodinium which is a gill parasite. Hence, we obtained a third group of fish and restarted the trial for the third time. This trial included 9 diets including the basal diet and diet for which Bright day (50, 100%), Hamlet300 (50, 100%), Soycomil (50%, 100%) and Expeller pressed soy (25 and 50%) were used to replace solvent extracted soybean meal on an equal protein basis. This trial was stocked 3/3/2023 and is expected to run for 8 weeks. At the conclusion of this trial, we will then need to sample the fish to evaluate possible nutrient retention differences as well as the effects of the various soy sources on gut health. For the histology work only the basal and 100% replacement dietary treatments will be evaluated. The distal portion of the gut will be excised, preserved and then sent to the veterinary school for sectioning and staining. Additionally, fish will need to be analyzed to determine protein levels which are required to look at apparent net protein retention. Unfortunately, this work will exceed the current end date. Consequently, we have asked for a no cast extension so that we have access to the funds to complete the analysis once the growth trial is completed.

The take home message from this work is that comparing a practical soy maximized diet to a commercial feed formulation commonly used with Florida pompano that there is a slight depression in growth. Thus, demonstrating that we are very close to using high soy feed formulations (based on solvent extracted soybean meal) for this species. Histological evaluations of the gut from these fish demonstrate that there is no enteritis (allergic response to soy) which means that depressions in performance are due either to the presence of antinutrients or inadequacies in nutrition of the feed. Hence, looking at other soy-products with lower levels of antinutrients may lead to better performance. This is what the second and ongoing component of the research is doing. We are looking at various advanced soy products and the effects in high soy feed formulations. Once completed, results of this work will add to the literature on use of soy based product in practical feed formulations for this species.

Updated September 1, 2023:
In the first objective of this work, we sought to compare our fishmeal free high soy practical diet (8% poultry meal, 10% corn protein concentrate, 55% soybean meal) with a commercial feed commonly used to rear the Florida pompano. Thus, providing validation of the high soy open feed formulation that we use in research. This work was conducted in two independent systems (Referred to as Trials 1 and 2 in attached tables). Juvenil fish (23.2g) were reared in our smaller tank (800 L) recirculation system using six replicates for each diet. At the conclusion of the 6-week growth final weight (55.2 and 60.4g, P=0.076), percent weight gain (138.8% and 160.0%; P=0.006) and FCR (2.05, 1.86; P= 0.015) for fish reared on the experimental feed were poorer but not necessarily significantly than that of fish reared on the commercial feed. A similar response was observed in our large tanks system (3,000 L) which utilized three replicate groups of larger fish with an initial mean weight of 38.1g. At the conclusion of the 8-week growth trial final weights (125g and 137g; P-value 0.3318), percent weight gain (226.1% and 269.2 %; P = 0.284) and FCR (2.05 and 1.89; p= 0.577) were not statistically different. In both cases, performance was lower, albeit not necessarily significantly (P < 0.05) in fish offered the experimental diet as compared to the commercial feed. This would indicate that although we were very close in terms of performance, we still need to optimize diets that have low levels of animal protein and high levels of plant-based proteins.

Samples of the distal gut were collected from fish reared in the 8-week trial. These samples were sectioned and stained. The lamina propria thickness and cellularity, as well as vacuolization and connective tissue thickness were evaluated. The slides were scored with no clear signs of enteritis noted. Based on the available data, enteritis is not present in high soy feed formulations which would indicate that other nutritional or antinutrients are responsible for any reduced performance.

The second component of this research was to evaluate various soybean sources. For this work different processed soybean meal products were sourced including: new variety; high protein low oligosaccharide solvent-extracted soybean meal (Benson Hill, St. Louis, MO, USA), fractionated soy protein concentrate (Soycomil, ADM Animal Nutrition, Quincy, IL, USA), fermented soybean meal (Hamlet HP300, Hamlet Protein Inc., Findlay, OH, USA), and modified expeller-pressed soybean meal (All Sustained LLC., Shrewsbury, USA) were used in diets. The idea was that the various processing methods would result in different antinutrient levels and potentially result in improved performance. The basal diet and 8 test diets were formulated (Tables 3 and 4, attached) to have equivalent levels of protein and lipids. Test diets were developed by replacing soybean meal (on an equal protein basis) with each product (Bright Day, Hamlet HP300, and Soycomil) at 50% and 100% replacement. Expeller-pressed soy was used only to replace 25% and 50% of the soybean meal. The growth trial was run at E. W. Shell Fisheries Center under controlled conditions in a recirculating aquaculture system. Four replicate groups of fish (4.82g mean weight) were assigned to each dietary treatment and reared over a 76-day culture period. At the conclusion of the growth trial (Tables 6 and 7, attached), there were no significant differences in terms of final weight (32.4 to 45.9g), percent gain (587.7 to 840.1%), FCR (1.88-2.19) or survival (86.6-97.5%). Although, the lack of statistically different results is disappointing. The lack of difference is, in my opinion, due to the variability of this fish as opposed to no biological response. In all replacement products, the fish were numerically larger or had higher weight gain as compared to those reared on solvent-extracted soybean meal, indicating that these products are likely of higher quality. For this work, we also preserved distal gut samples to confirm the histology of the initial trial. The samples have been processed, but histological quantification and qualitative scoring is ongoing. The results from this study will be compiled, presented at scientific meetings, and a paper will be developed from the data.

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The take-home message from this work is that comparing a practical soy-maximized diet to a commercial feed formulation commonly used with Florida pompano that there is a slight depression in growth. Thus, demonstrating that we are very close to using high soy feed formulations (based on solvent-extracted soybean meal) for this species. Histological evaluations of the gut from these fish are underway and preliminarily demonstrate that there is no enteritis (allergic response to soy). This likely means that the observed growth depressions in performance are due either to the presence of antinutrients or inadequacies in nutrition of the feed. Hence, looking at other soy products with lower levels of antinutrients may lead to better performance. This is what the second and ongoing component of the research is doing. We are looking at various advanced soy products and the effects in high soy feed formulations. Results of this work will add to the literature on use of soy-based product in practical feed formulations for this species.

The data will expand our understanding of nutritional limitations of soy-based diets for marine fish and demonstrate their use for extended growout of fish. Results will promote the use of soybean meal as a sustainable and cost-effective protein source in marine fish feeds. Additionally, if new varieties are going to be marketed as high value products, we need data with high values specie for which the use of specialty ingredients makes sense.