Application of laboratory methods for understanding fish responses to black soldier fly (Hermetia illucens) based diets

A major challenge for development of sustainable aquafeeds is its dependence on fish meal and fish oil. Replacement with more sustainable, nutritious and safe ingredients is now a priority. Over the last years, among several alternatives proposed, insects have received great attention as possible candidates. In particular, the black soldier fly (Hermetia illucens; BSF) represents a concrete example of how the circular economy concept can be applied to fish culture, providing a valuable biomass rich in fat and protein valorising organic by-products. In the last decade, several studies have been published about the use of different BSF dietary inclusion levels for various fish species including experimental models. Varying and encouraging results have been obtained in this research field using a plethora of laboratory methodological approaches that can be applied and coupled to obtain a comprehensive view of the BSF-based diets effects on fish physiology, health, and quality. The present review aims to explore some of the most promising laboratory approaches like histology, infrared spectroscopy, gut microbiome sequencing, molecular biology, fish fillets’ physico-chemical and sensory properties, essential for a better understanding of fish welfare and fillet quality, when BSF is used as aquafeed ingredient. In particular, great importance has been given to European finfish species and experimental models.publishedVersio

Hermetia illucens and Poultry by-Product Meals as Alternatives to Plant Protein Sources in Gilthead Seabream (Sparus aurata) Diet: A Multidisciplinary Study on Fish Gut Status

The attempt to replace marine-derived ingredients for aquafeed formulation with plant-derived ones has met some limitations due to their negative side effects on many fish species. In this context, finding new, sustainable ingredients able to promote fish welfare is currently under exploration. In the present study, poultry by-products and Hermetia illucens meal were used to replace the vegetable protein fraction in diets totally deprived of fish meal intended for gilthead seabream (Sparus aurata). After a 12-week feeding trial, a multidisciplinary approach including histological, molecular, and spectroscopic techniques was adopted to investigate intestine and liver responses to the different dietary formulations. Regardless of the alternative ingredient used, the reduction in dietary vegetable proteins resulted in a lower incidence of intestine histological alterations and inflammatory responses. In addition, the dietary inclusion of insect meal positively affected the reduction in the molecular inflammatory markers analyzed. Spectroscopic analyses showed that poultry by-product meal improved lipid absorption in the intestine, while insect meal induced increased liver lipid deposition in fish. The results obtained demonstrated that both poultry by-products and H. illucens meal can successfully be used to replace plant-derived ingredients in diets for gilthead seabream, promoting healthy aquaculture

Effects of Graded Dietary Inclusion Level of Full-Fat Hermetia illucens Prepupae Meal in Practical Diets for Rainbow Trout (Oncorhynchus mykiss)

This study investigated the effects of dietary inclusion levels of full-fat Hermetia illucens prepupae meal (H) on growth and gastrointestinal integrity in rainbow trout (Oncorhynchus mykiss). A 98-day study was conducted using triplicate groups of trout (initial body weight, 137 \ub1 10.5 g) kept in 1-m3 tanks in a flow-through well water system. Three dietary treatments were prepared: one based on fishmeal and purified protein-rich vegetable ingredients (H0), and two experimental diets including graded levels of H meal (25% and 50%, referred to as H25 and H50, respectively). At the end of the feeding trial, no differences were observed in growth performance and plasma metabolite levels, with the biometric data confirmed by the liver expression of the genes involved in somatic growth regulation (igf1 and mstn1a). In the H50 group, a three-fold up regulation of liver hsp70 was observed. An activation of the stress/immune response (il-10, tnf-\u3b1, and tlr-5) was observed in medium intestine in the H25 and H50 groups (p < 0.05) together with a villi length reduction detected through histological analyses. Liver histology and Fourier Transform Infrared Imaging (FTIRI) spectroscopy highlighted an increase in lipid deposition. These findings suggest that caution should be taken into account when 50% replacement of conventional ingredients with H is selected

Appetite Regulation, Growth Performances and Fish Quality Are Modulated by Alternative Dietary Protein Ingredients in Gilthead Sea Bream (Sparus aurata) Culture

By answering the need for increasing sustainability in aquaculture, the present study aimed to compare growth, gene expression involved in appetite regulation, physical characteristics, and chemical composition of Sparus aurata fed alternative protein sources. Fish were fed ten iso-proteic, iso-lipidic, and isoenergetic diets: a vegetable-based (CV) and a marine ingredient-rich (CF) diet were set as control diets. The others were prepared by replacing graded levels (10, 20 or 40%) of the vegetable proteins in the CV with proteins from a commercial defatted Hermetia illucens pupae meal (H), poultry by-product meal (PBM) singly (H10, H20, H40, P20, P40) or in combination (H10P30), red swamp crayfish meal (RC10) and from a blend (2:1, w:w) of Tisochrysis lutea and Tetraselmis suecica (MA10) dried biomasses. The increase in ghre gene expression observed in MA10 fed fish matched with increased feed intake and increased feed conversion ratio. Besides, the MA10 diet conferred a lighter aspect to the fish skin (p < 0.05) than the others. Overall, no detrimental effects of H, PBM, and RC meal included in the diets were observed, and fish fatty acid profile resulted as comparable among these groups and CV, thus demonstrating the possibility to introduce H, PBM, and RC in partial replacement of vegetable proteins in the diet for Sparus aurata

Effects of Di-Isononyl Phthalate (DiNP) on Follicular Atresia in Zebrafish Ovary

Di-isononyl phthalate (DiNP) is a plasticizer reported to elicit hormone-like activity and disrupt metabolism and reproduction in fish and other vertebrates. In general, phthalates have been used at high concentrations beyond reported environmental levels to assess their adverse effects on fish gonadal physiology. The present study exposed adult female zebrafish to a wide range of DiNP concentrations [0.42 µg L−1 (10−9 M), 4.2 µg L−1 (10−8 M), and 42 µg L−1 (10−7 M)] for 21 days. We evaluated gene expression profiles related to apoptosis, autophagy, and oxidative stress; DNA fragmentation (TUNEL assay: terminal deoxynucleotidyl transferase dUTP nick end labeling) and caspase activity (CAS3) were also examined. Exposure to 0.42 and 4.2 µg L−1 upregulated the genes coding for tnfa and baxa, sod1, prkaa1, respectively. CAS3 immunohistochemistry revealed a higher number of positive vitellogenic oocytes in ovaries exposed to 0.42 µg L−1. Subsequently, we examined the relationship between CAS3 signaling and DNA fragmentation. Accordingly, DNA fragmentation was observed in vitellogenic follicles of fish exposed to 0.42 and 4.2 μg L−1. Our results demonstrate that follicular atresia can occur after exposure to environmental levels of DiNP for 21 days, which may adversely affect the reproductive performance of female zebrafish in a non-monotonic manner.Fil: Godoi, Filipe G. Andrade. Università Politecnica dele Marche; Italia. Universidade de Sao Paulo; BrasilFil: Forner Piquer, Isabel. Università Politecnica dele Marche; ItaliaFil: Randazzo, Basilio. Università Politecnica dele Marche; ItaliaFil: Habibi, Hamid R.. University of Calgary; CanadáFil: Lo Nostro, Fabiana Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Biodiversidad y Biología Experimental y Aplicada. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biodiversidad y Biología Experimental y Aplicada; ArgentinaFil: Moreira, Renata Guimarães. Universidade de Sao Paulo; BrasilFil: Carnevali, Oliana. Università Politecnica dele Marche; Italia. Consorzio Interuniversitario di Biosistemi e Biostrutture; Itali

Replacing Maize Grain with Ancient Wheat Lines By-Products in Organic Laying Hens' Diet Affects Intestinal Morphology and Enzymatic Activity

The effects of replacement of maize grain with ancient wheat by-products on intestinal morphometry and enzymatic activity in laying hens was studied. Eighty hens were divided into two groups (40 each, 8 replicates, 5 hens/replicate) fed two isoproteic and isoenergetic diets. In the treated group, part of the maize was replaced by a mix of ancient grains (AGs) middling, in a 50:50 ratio of Triticum aestivum L. var. spelta (spelt) and Triticum durum dicoccum L. (emmer wheat). The AG diet affected the weight of all the large intestine tracts, decreasing the weight of caeca (p < 0.01) and increasing those of colon (p < 0.01), rectum and cloaca (p < 0.05). Villus height in the AG group was higher (p < 0.01) than the control for the duodenum and jejunum, while for the ileum, the control group showed the highest values (p < 0.01). The submucosa thickness was higher (p < 0.01) in the control group for the duodenum and ileum, while the jejunum for the AG group showed the highest (p < 0.05) submucosa thickness. The crypts depth was higher (p < 0.01) in the control group for the duodenum and ileum. Enzyme activity was enhanced by AGs (p < 0.01) in the duodenum. Regarding the jejunum, sucrase-isomaltase and alkaline phosphatase had higher activity (p < 0.05 and p < 0.01, respectively) in the AG group. In the ileum, sucrase-isomaltase showed higher activity (p < 0.01) in the control group, while alkaline phosphatase showed the highest values (p < 0.05) in the AG group. Overall, results suggested that the dietary inclusion of AGs exerted positive effects in hens, showing an improved intestinal function

Replacing Maize Grain with Ancient Wheat Lines By-Products in Organic Laying Hens’ Diet Affects Intestinal Morphology and Enzymatic Activity

The effects of replacement of maize grain with ancient wheat by-products on intestinal morphometry and enzymatic activity in laying hens was studied. Eighty hens were divided into two groups (40 each, 8 replicates, 5 hens/replicate) fed two isoproteic and isoenergetic diets. In the treated group, part of the maize was replaced by a mix of ancient grains (AGs) middling, in a 50:50 ratio of Triticum aestivum L. var. spelta (spelt) and Triticum durum dicoccum L. (emmer wheat). The AG diet affected the weight of all the large intestine tracts, decreasing the weight of caeca (p < 0.01) and increasing those of colon (p < 0.01), rectum and cloaca (p < 0.05). Villus height in the AG group was higher (p < 0.01) than the control for the duodenum and jejunum, while for the ileum, the control group showed the highest values (p < 0.01). The submucosa thickness was higher (p < 0.01) in the control group for the duodenum and ileum, while the jejunum for the AG group showed the highest (p < 0.05) submucosa thickness. The crypts depth was higher (p < 0.01) in the control group for the duodenum and ileum. Enzyme activity was enhanced by AGs (p < 0.01) in the duodenum. Regarding the jejunum, sucrase-isomaltase and alkaline phosphatase had higher activity (p < 0.05 and p < 0.01, respectively) in the AG group. In the ileum, sucrase-isomaltase showed higher activity (p < 0.01) in the control group, while alkaline phosphatase showed the highest values (p < 0.05) in the AG group. Overall, results suggested that the dietary inclusion of AGs exerted positive effects in hens, showing an improved intestinal function

Towards the Identification of a Suitable Commercial Diet for Carpione (Salmo carpio, Linnaeus 1758): A Multidisciplinary Study on Fish Performances, Animal Welfare and Quality Traits

SIMPLE SUMMARY: Carpione (Salmo carpio, Linnaeus 1758) is an endangered precious endemism of Lake Garda (Northern Italy), the largest Italian lake. To date, several bottlenecks about its culture remain unsolved, including the identification of a proper growth-out diet. In the present study, four different grossly isolipidic, isoproteic, and isoenergetic diets containing ingredients from different origins were used for S. carpio culture. Specifically, a diet largely based on marine ingredients, and currently used for carpione farming, was used as control. Three other diets were formulated in order to include relevant percentages of vegetable ingredients or processed animal proteins (at two different inclusion levels). After a three-month feeding trial, fish zootechnical performances, welfare, and flesh quality were evaluated through a multidisciplinary approach, including histology, gene expression, chemical analysis, and Fourier transform infrared spectroscopy (FTIR). This study provided the first insights on carpione physiological responses to different commercial dietary formulations. ABSTRACT: Carpione (Salmo carpio, Linnaeus 1758) is an endangered precious endemism of Lake Garda (Northern Italy), the largest Italian lake. To date, several bottlenecks about its culture remain unsolved, including the identification of a proper growth-out diet. The aim of the present study was to test four different grossly isolipidic, isoproteic, and isoenergetic diets in which the main ingredients had a different origin. Specifically, a diet currently used by local farmers for carpione culture, largely based on marine ingredients, was used as control (CTRL), while the other three diets were formulated by partially replacing marine ingredients with plant ones (VEG) or with different percentages of processed animal proteins (PAP1 and PAP2). The feeding trial was run in triplicate, over a three-month period. No significant differences in growth performance among the experimental groups were observed. However, remarkable histological alterations and inflammatory markers upregulation were observed in VEG group, while PAP inclusion played a role in attenuating inflammation and improving nutrient uptake. Fillet analyses highlighted significant differences in marketable traits and flesh fatty acid composition among the experimental groups, including the reduction of polyunsaturated fatty acids related to PAPs inclusion. In conclusion, PAPs used in the present study promoted S. carpio gut health and absorption capacity, while further studies are required to maintain proper quality traits of the final product