Protein hydrolysate and oil from fish waste reveal potential as dog food ingredients.

The increased fish consumption by the growing human population in the world translates into an increase in fish waste. The reintroduction of these fish by-products into food and feed chains presents economic benefits and contributes to counteracting their negative environmental impact. Under this context, the present study aimed to evaluate the effects of the dietary inclusion of fish hydrolysate and oil obtained from fish waste (experimental diet) in substitution of shrimp hydrolysate and salmon oil (control diet) mainly imported from third countries on palatability, apparent total tract digestibility, fecal characteristics and metabolites, blood fatty acid profile, flatulence, and coat quality of adult dogs. A two-bowl test was performed to evaluate palatability by the pairwise comparison between the two diets. A feeding trial was conducted according to a crossover design with two diets (control and experimental diets), six adult Beagle dogs per diet, and two periods of 6 weeks each. The replacement of shrimp hydrolysate and salmon oil with fish hydrolysate and oil did not affect the first diet approach and taste, as well as the intake ratio. Generally, the digestibility of dry matter, nutrients, and energy was not affected by diet, but the intake of digestible crude protein (CP) and ether extract was higher, respectively, with the control and the experimental diet. The higher intake of eicosapentaenoic acid and docosahexaenoic acid with the experimental diet was reflected in a higher content of these long-chain polyunsaturated fatty acids and the omega-3 index of red blood cells, but it did not affect coat quality. The significantly higher intake of digestible CP with the control diet might have contributed to the higher fecal ammonia-N and valerate concentrations. Daily fecal output and characteristics were similar between diets. Overall, results suggest that fish hydrolysate and oil from the agrifood industry might constitute sustainable functional ingredients for dog feeding while adding value for wild fisheries, aquaculture, and fish farming under a circular economy approach and reducing dependence on imports from third countries with a high carbon footprint.

Environmental conditions affect the nutritive value and alkaloid profiles of Lupinus forage: Opportunities and threats for sustainable ruminant systems.

The identification of crops that simultaneously contribute to the global protein supply and mitigate the effects of climate change is an urgent matter. Lupins are well adapted to nutrient-poor or contaminated soils, tolerate various abiotic stresses, and present relevant traits for acting as ecosystem engineers. Lupins are best studied for their seeds, but their full foraging potential needs further evaluation. This study evaluated the effects of location and sowing date on forage production, proximate composition, and the detailed mineral and alkaloid profiles of three species of Lupinus (L. albus cv. Estoril, L. angustifolius cv. Tango, and L. luteus cv. Cardiga). Sowing date and location and their interaction with the plant species significantly affected the vast majority of measured parameters, emphasizing the effects of climate and soil conditions on these crops. The relatively high crude protein and in vitro digestibility support the potential of the lupin species studied as sustainable forage protein sources in diets for ruminant animals. The content of individual essential macro and trace elements was below the maximum tolerable levels for cattle and sheep. Lupanine, smipine, and sparteine were the most abundant quinolizidine alkaloids in L. albus cv. Estoril, lupanine, and sparteine in L. angustifolius cv. Tango, and lupinine, gramine, ammodendrine, and sparteine in L. luteus cv. Cardiga. Based on the maximum tolerable levels of total quinolizidine alkaloid intake, the dietary inclusion of forages of L. albus cv. Estoril and L. angustifolius cv. Tango does not pose a risk to the animals, but the high alkaloid content of L. luteus cv. Cardiga may compromise its utilization at high levels in the diet. Overall, the results reveal a high potential for lupins as protein forage sources well adapted to temperate regions and soils with lower fertility, with a relevant impact on livestock sustainability in a climate change era.

Agronomic, Nutritional Traits, and Alkaloids of Lupinus albus, Lupinus angustifolius and Lupinus luteus Genotypes: Effect of Sowing Dates and Locations.

Lupins (Lupinus spp.) are legumes with high relevance for the sustainability of agricultural systems as they improve the soil quality, namely, through the fixation of atmospheric nitrogen, and have good adaptability to different climates and soil conditions. Besides, they possess high nutritive value, especially due to the high protein content of the seeds. Nevertheless, the plants' productivity and metabolism can be influenced by the genotype, the edaphoclimatic conditions, and the sowing practices. In this work, the effect of edaphoclimatic conditions and sowing dates on the productivity, nutritional factors, and alkaloids of the seeds of L. albus cv. Estoril, L. angustifolius cv. Tango, and L. luteus cv. Cardiga was evaluated. High variability in the seeds and protein productions, nutritional traits, and alkaloid content related to the species was observed, along with a significant effect of the location. Lupinus albus cv. Estoril showed a good compromise between productivity and low alkaloid content, being an interesting genotype for food and feed use in the conditions of this trial.

Ruminal inocula with distinct fermentation profiles differentially affect the in vitro fermentation pattern of a commercial algal blend.

The in vitro rumen batch technique is widely used for screening novel feed sources; however, it remains unclear to what extent the in vitro fermentability of non-conventional feed sources is affected by non-adapted ruminal inocula. Thus, in this study, we evaluated the effects of distinct ruminal inocula on the in vitro fermentation parameters of a sustainable non-conventional feed, a commercially available algal blend composed of microalgae (Chlorella vulgaris and Nannochloropsis oceanica) and seaweeds (Ulva sp. and Gracilaria gracilis). First, four late-lactation Holstein cows were fed four forage-based diets varying only in the proportions of basal forage (100% corn silage, 70% corn silage and 30% haylage, 30% corn silage and 70% haylage, and 100% haylage) in a 4 × 4 Latin square design with the last square omitted. After 3 weeks of adaptation, haylage-based diets resulted in ruminal fermentation parameters distinct from those promoted by corn silage-based diets, as reflected in increased pH, ammonia-N contents, and acetate proportions. Individual ruminal fluids derived from each of the four diets were further used as inocula in in vitro incubations. Here, a 1:1 mixture of corn silage and haylage was supplemented with 0, 5, 10, or 15% algal blend and incubated with each inoculum for 24 h in a 4 × 4 factorial design. Total gas and methane production decreased with inocula from cows fed haylage-based diets and with increasing algal blend supplementation levels. The fermentation pH increased and the ammonia-N contents decreased with inocula from cows fed haylage-based diets; however, these parameters were not affected by algal blend inclusion levels. The interaction between the ruminal inoculum source and the algal blend supplementation level affected the total volatile fatty acids (VFA) and the proportions of most individual VFA. Total VFA production decreased with increasing algal supplementation levels, particularly with inocula from cows fed 30% corn silage and 70% haylage; the acetate, propionate, and valerate proportions were only affected by algal blend levels under incubation with 100% corn silage inocula. Overall, our findings highlight the importance of the ruminal inoculum source when assessing the fermentability of non-conventional feed as well as the potential of the algal blend as a natural modulator of ruminal fermentation.

Squid meal and shrimp hydrolysate as novel protein sources for dog food.

The world's growing pet population is raising sustainability and environmental concerns for the petfood industry. Protein-rich marine by-products might contribute to mitigating negative environmental effects, decreasing waste, and improving economic efficiency. The present study evaluated two marine by-products, squid meal and shrimp hydrolysate, as novel protein sources for dog feeding. Along with the analysis of chemical composition and antioxidant activity, palatability was evaluated by comparing a commercial diet (basal diet) and diets with the inclusion of 150 g kg-1 of squid meal or shrimp hydrolysate using 12 Beagle dogs (2.2 ± 0.03 years). Two in vivo digestibility trials were conducted with six dogs, three experimental periods (10 days each) and three dietary inclusion levels (50, 100 and 150 g kg-1) of squid meal or shrimp hydrolysate in place of the basal diet to evaluate effects of inclusion level on apparent total tract digestibility (ATTD), metabolizable energy content, fecal characteristics, metabolites, and microbiota. Both protein sources presented higher protein and methionine contents than ingredients traditionally used in dog food formulation. Shrimp hydrolysate showed higher antioxidant activity than squid meal. First approach and taste were not affected by the inclusion of protein sources, but animals showed a preference for the basal diet. Effects on nutrient intake reflected the chemical composition of diets, and fecal output and characteristics were not affected by the increasing inclusion levels of both protein sources. The higher ATTD of dry matter, most nutrients and energy of diets with the inclusion of both by-products when compared to the basal diet, suggests their potential to be included in highly digestible diets for dogs. Although not affected by the inclusion level of protein sources, when compared to the basal diet, the inclusion of squid meal decreased butyrate concentration and shrimp hydrolysate increased all volatile fatty acids, except butyrate. Fecal microbiota was not affected by squid meal inclusion, whereas inclusion levels of shrimp hydrolysate significantly affected abundances of Oscillosperaceae (UCG-005), Firmicutes and Lactobacillus. Overall, results suggest that squid meal and shrimp hydrolysate constitute novel and promising protein sources for dog food, but further research is needed to fully evaluate their functional value.

Effects of microalgae as dietary supplement on palatability, digestibility, fecal metabolites, and microbiota in healthy dogs.

The current trend of dog owners increasingly favoring the functional value of food to assure preventive health and wellbeing of their pets has been raising the interest in microalgae as natural additives with bioactive properties. However, scientific studies addressing the effects of microalgae supplementation in diets for dogs are scarce. This study aimed to evaluate the effects of dietary supplementation with three microalgae species (Chlorella vulgaris, Nannochloropsis oceanica, and Tetradesmus obliquus) on diet palatability, total tract digestibility, metabolizable energy content, fecal metabolites and microbiota of dogs. Twelve adult Beagle dogs were used in three two-bowl tests to compare the palatability of a commercial complete diet for adult dogs without (reference diet) and with 1.5% supplementation of each microalgae. From the results obtained, three digestibility trials were performed according to a replicated Latin square 3 × 3, with six adult Beagle dogs, three experimental periods of 10 days each, and three dietary supplementation levels of microalgae (0.5, 1.0, and 1.5%). In each trial, effects of microalgae supplementation levels on total tract digestibility, metabolizable energy content, fecal metabolites and microbiota of dogs were evaluated. First diet approached or tasted was not significantly affected by microalgae inclusion, but dogs showed a preference for the reference diet over the diets with 1.5% inclusion of C. vulgaris and N. oceanica, no difference being observed with 1.5% T. obliquus. In all digestibility trials, dietary supplementation with microalgae up to 1.5% did not greatly affected the dietary chemical composition and kept unaffected food intake, fecal output and metabolites, and digestibility of nutrients and energy. Compared with the reference diet, supplementation with C. vulgaris increased protein digestibility. Fecal characteristics and metabolites were affected by microalgae supplementation, being the effects dependent on the species. Fecal microbiota composition of dogs fed with microalgae-supplemented diets was modified by promoting the beneficial Turicibacter and Peptococcus genera associated with gut health and activation of the immune system. Overall, the results support C. vulgaris, N. oceanica, and T. obliquus as sustainable functional supplements that potentially enhance gastrointestinal health of dogs through the selective stimulation of microbiota without detrimental effects on food intake and digestibility.

Upcycling post-harvest biomass residues from native European Lupinus species: from straws and pod shells production to nutritive value and alkaloids content for ruminant animals.

The production of Lupinus seeds for food and feed is increasing worldwide, which results in large amounts of post-harvest biomass residues, considered of low value and left in the field to be burned or incorporated in the soil. To valorize these agricultural wastes, this work aimed to assess their potential as an alternative feed for ruminants. Thus, the production yield, nutritive value, and alkaloid content of straws and pod shells from three native European Lupinus species, L. albus 'Estoril' (white), L. angustifolius 'Tango' (narrow-leafed), and L. luteus 'Cardiga' (yellow), cultivated in two locations, were evaluated. The dry matter (DM) yield of straws and pod shells were the highest for L. albus 'Estoril' (4.10 t ha-1) and the lowest for L. angustifolius 'Tango' (1.78 t ha-1), suggesting a poor adaptation of narrow-leafed lupin to the particularly dry and warm agronomic year. Despite species-specific differences, lupin biomass residues presented higher crude protein (53.0-68.9 g kg-1 DM) and lignin (103-111 g kg-1 DM) content than cereal straws usually used in ruminant feeding, thus resulting in higher metabolizable energy (6.43-6.58 MJ kg-1 DM) content. In vitro digestibility was similar among lupin species (47.7-50.6%) and higher in pod shells (53.7%) than in straws (44.6%). Lupinus albus 'Estoril' and L. luteus 'Cardiga' presented considerable amounts of alkaloids in straws (23.9 and 119 mg kg-1 DM) and pod shells (20.5 and 298 mg kg-1 DM), while no alkaloids were detected in L. angustifolius 'Tango' biomass residues. Considering the combined production of straw and pod shells per lupin species, it is anticipated that lupin biomass residues produced per ha can fulfill 85% of the energy and nearly 50% of protein requirements of a flock of 4 to 9 dry and mid-pregnancy sheep with 50 kg body weight for one year. No negative effects on small (ovine) and large (bovine) ruminant species due to alkaloids are expected, even if biomass residues are consumed at up to 85% DM intake. The large production yield along with its nutritive value unveils the potential of lupin biomass residues valorization as alternative fodder for ruminants, promoting sustainability under a circular economy approach.

A commercial blend of macroalgae and microalgae promotes digestibility, growth performance, and muscle nutritional value of European seabass (Dicentrarchus labrax L.) juveniles.

Algae can leverage aquaculture sustainability and improve the nutritional and functional value of fish for human consumption, but may pose challenges to carnivorous fish. This study aimed to evaluate the potential of a commercial blend of macroalgae (Ulva sp. and Gracilaria gracilis) and microalgae (Chlorella vulgaris and Nannochloropsis oceanica) in a plant-based diet up to 6% (dry matter basis) on digestibility, gut integrity, nutrient utilization, growth performance, and muscle nutritional value of European seabass juveniles. Fish (11.3 ± 2.70 g) were fed with isoproteic, isolipidic, and isoenergetic diets: (i) a commercial-type plant-based diet with moderate fishmeal (125 g kg-1 DM basis) and without algae blend (control diet; Algae0), (ii) the control diet with 2% algae blend (Algae2), (iii) the control diet with 4% algae blend (Algae4), and (iv) the control diet with 6% algae blend (Algae6) for 12 weeks. The digestibility of experimental diets was assessed in a parallel study after 20 days. Results showed that most nutrients and energy apparent digestibility coefficients were promoted by algae blend supplementation, with a concomitant increase in lipid and energy retention efficiencies. Growth performance was significantly promoted by the algae blend, the final body weight of fish fed Algae6 being 70% higher than that of fish fed Algae0 after 12 weeks, reflecting up to 20% higher feed intake of algae-fed fish and the enhanced anterior intestinal absorption area (up to 45%). Whole-body and muscle lipid contents were increased with dietary algae supplementation levels by up to 1.79 and 1.74 folds in Algae 6 compared to Algae0, respectively. Even though the proportion of polyunsaturated fatty acids was reduced, the content of EPA and DHA in the muscle of algae-fed fish increased by nearly 43% compared to Algae0. The skin and filet color of juvenile European seabass were significantly affected by the dietary inclusion of the algae blend, but changes were small in the case of muscle, meeting the preference of consumers. Overall results highlight the beneficial effects of the commercial algae blend (Algaessence®) supplementation in plant-based diets for European seabass juveniles, but feeding trials up to commercial-size fish are needed to fully assess its potential.

Insights from the yield, protein production, and detailed alkaloid composition of white (Lupinus albus), narrow-leafed (Lupinus angustifolius), and yellow (Lupinus luteus) lupin cultivars in the Mediterranean region.

Introduction: Lupins and other legumes have been considered as alternative plant-based protein sources to soybeans for both humans and livestock. Furthermore, they can contribute to more sustainable agricultural systems. The productivity and chemical composition of legumes is highly variable between species, cultivars, and with the edaphoclimatic conditions. Methods: This work evaluated the adaptability of seven Lupinus cultivars in two different sowing locations, during two consecutive years, through the characterization of their seed, as a means of investigating their suitability to be used as a source of food and/or feed. Results and discussion: Lupinus angustifolius cv. Tango and Lupinus luteus cv. Acos were the most stable genotypes across the environments when considering the seed and protein production, while L. luteus cv. Alburquerque and L. luteus cv. Mister showed less variation in the total alkaloid content across the environments. The edaphoclimatic conditions affected seed and protein yields, as higher rainfall resulted in high productivity. The lower temperatures observed in the first year at both locations caused a reduction in the production of alkaloids in L. luteus cv. Acos and Cardiga. Due to the high alkaloid content of some of the studied cultivars their use as food or feed can pose some safety concerns. However, these cultivars can have high levels of resistance to herbivore and insect attacks, which can be of the utmost importance for the use of these crops for recovering poor or exhausted soils.

Nutritional Composition and Untargeted Metabolomics Reveal the Potential of Tetradesmus obliquus, Chlorella vulgaris and Nannochloropsis oceanica as Valuable Nutrient Sources for Dogs.

The growing pet population is questioning the sustainability of the pet food system. Although microalgae may constitute a more sustainable food resource, the assessment of their potential for canine diets is almost non-existent. The present study aimed to evaluate the potential of three microalgae species (Tetradesmus obliquus, Chlorella vulgaris and Nannochloropsis oceanica) grown locally in industrial photobioreactors as alternative food resources for dogs. A detailed characterization of their nutritional composition and metabolomic profile was carried out and related to the nutritional requirements of dogs. Overall, the essential amino acid content exceeded the amounts required for dogs at all life stages, except methionine and cysteine. The three microalgae were deficient in linoleic acid, N. oceanica presented a linolenic acid content below requirements and T. obliquus and C. vulgaris were deficient in arachidonic and eicosapentaenoic acids. The fiber was mainly composed of insoluble dietary fiber. The mineral profile varied greatly with the microalgae species, demonstrating their different potential for dog feeding. Untargeted metabolomics highlighted glycolipids, glycerolipids and phospholipids as the most discriminating compounds between microalgae species. Overall, the results support the potential of T. obliquus, C. vulgaris and N. oceanica as valuable macro- and micro-nutrients sources for dog feeding.

Ammonia and greenhouse emissions from cow's excreta are affected by feeding system, stage of lactation and sampling time.

Decomposition of dairy cows' excreta on housing floor leads to ammonia and greenhouse gases production, yet factors affecting total emissions have not been fully disclosed. This work aimed to assess the impact of lactation stage, feeding system and sampling time on gaseous emission potential of cow's faeces and urine in laboratory chambers systems. Individual faeces and urine were collected from two groups of four cows, at peak and post peak lactation, from three commercial farms with distinct feeding systems: total mixed ration (TMR), total mixed ration plus concentrate at robot (TMR + robot), and total mixed ration plus concentrate in automatic feeders (TMR + AF). Samples were collected before a.m. (T8h), at middle day (T12h), and before p.m. (T17h) milking. In a laboratory chambers system, faeces and urine were mixed in a ratio of 1.7:1, and ammonia and greenhouse gases emissions were monitored during 48-h. Cumulative N-N2O emissions were the highest in TMR + robot system, post peak cows and sampling time T17h. An interaction between stage of lactation and sampling time was detected for N-NH3 and N-N2O (g/kg organic soluble N) emissions. Post peak cows also produced the highest cumulative N-NH3 emissions. Overall results contribute for the identification of specific on-farm strategies to reduce gaseous emissions from cows' excreta.

Ecotoxicity of Recycled Aggregates: Application of a Prediction Methodology.

Due to environmental concerns, the search for sustainable construction solutions has been increasing over the years. This global concern is creating a trend in the use of recycled aggregates resulting from construction and demolition wastes from different sources. In addition to their physical and mechanical properties, it is important to analyse their ecotoxicological risk to determine whether their leachates might be an issue. To assess ecotoxicity, biological tests should be performed for different trophic levels. This type of test is expensive and needs a high level of expertise, which leads to a lack of studies on recycled aggregates including ecotoxicity analysis. This paper presents a set of predictive ecotoxicity results based on the published studies on recycled aggregates. These results are the outcome of applying an innovative methodology previously developed and validated by the authors aiming to foresee the ecotoxicological fate of building materials' constituents and products. The application of this methodology enables the classification of a recycled aggregate product as safe or unsafe in terms of ecotoxicity risk, while keeping biological testing to a minimum.

A Novel Approach for Monitoring the Volatile Metabolome in Biological Samples from Ruminants through Miniaturized Liquid-Liquid Extraction and Multiclass Gas Chromatography Analysis.

A straightforward and versatile methodology for the extraction of volatile metabolites in biological samples from ruminants for gas chromatography analysis is proposed. The methodology was applied in the determination of multiclass metabolites (short-chain fatty acids, aldehydes, alcohols, ketones, esters, phenols, and sulfides) in different analytical matrices (rumen fluid, urine, and feces) collected from Holstein cows. The 24 multiclass volatile metabolites reported in the different biological samples and their respective concentrations were critically discussed in the context of digestive physiology. Most detected compounds are derived from the rumen and lower gut fermentation of carbohydrates, proteins, and lipids or their metabolism, being consistent with the prior state of the art. The proposed method also takes advantage of the already existing tools in animal nutrition laboratories, providing a novel methodological ground that can generate relevant bioanalytical information with a significant impact on ruminant's nutritional studies.

Effects of Zinc Source and Enzyme Addition on the Fecal Microbiota of Dogs.

Supplemental zinc from organic sources has been suggested to be more bioavailable than inorganic ones for dog foods. However, the bioavailability of zinc might be affected by dietary constituents such as phytates. The present study aimed to evaluate the effects of two zinc sources (zinc sulfate and zinc proteinate) and the addition of a multi-enzymatic complex from the solid-state fermentation of Aspergillus niger on end-products of fecal fermentation and fecal microbiota of adult Beagles fed a high-phytate diet. The experimental design consisted of three 4 × 4 Latin Squares with a 2 × 2 factorial arrangement of treatments (n = 12 Beagles), with four periods and four diets: zinc sulfate without (IZ) or with (IZ +) enzyme addition, and zinc proteinate without (OZ) or with (OZ +) enzyme addition. Enzyme addition significantly affected Faith's phylogenetic diversity index, whereas zinc source did not affect either beta or alpha diversity measures. Linear discriminant analysis effect size detected nine taxa as markers for organic zinc, 18 for inorganic source, and none for enzyme addition. However, with the use of a negative binomial generalized linear model, further effects were observed. Organic zinc was associated with a significantly higher abundance of Firmicutes and lower Proteobacteria and Bacteroidetes, although at a genus level, the response varied. The DNA abundance of Clostridium cluster I, Clostridium cluster XIV, Campylobacter spp., Ruminococcaceae, Turicibacter, and Blautia was significantly higher in dogs fed IZ and IZ + diets. Higher abundance of genus Lactobacillus was observed in dogs fed enzyme-supplemented diets. End-products of fecal fermentation were not affected by zinc source or enzymes. An increase in some taxa of the phyla Actinobacteria and Firmicutes was observed in feces of dogs fed organic zinc with enzyme addition but not with inorganic zinc. This study fills a gap in knowledge regarding the effect of zinc source and enzyme addition on the fecal microbiota of dogs. An association of zinc bioavailability and bacteria abundance is suggested, but the implications for the host (dog) are not clear. Further studies are required to unveil the effects of the interaction between zinc sources and enzyme addition on the fecal microbial community.

Energy: Protein Ratio in Ruminants: Insights from the Intragastric Infusion Technique.

Studies on energy:protein ratio in ruminants are constrained by rumen fermentation since it governs nutrient metabolism and the ratio of energy:protein yielding nutrients available for absorption. By circumventing rumen fermentation, the total intragastric infusion technique (IIT) allowed objective quantification of maintenance energy and protein requirements, volatile fatty acid utilisation efficiency, efficiency of energy utilisation for maintenance (Km) and growth (Kf) and the origin of N retention responses to independent variation of energy and protein intake. This review outlines the key IIT findings and whether they are reflected in current feeding systems with implications for different production systems worldwide. Maintenance energy requirements are similar to those derived from comparative slaughter but maintenance N requirements are significantly lower. No differences in utilisation efficiency exist between acetic, propionic and butyric acids. At low energy intakes, endogenous energy reserves are utilised to retain amino acids and fuel substantial tissue protein gains. The use of fasting metabolism to measure the utilisation of nutritionally balanced diets is questioned since it is a glucose-deficient state. Inter-species differences in glucose metabolism appear to exist, suggesting that glucose requirements may be higher in cattle than sheep. The difficulty in predicting nutrient requirements, particularly protein, with any one technique is highlighted.

Validation of a Simple HPLC-Based Method for Lysine Quantification for Ruminant Nutrition.

Robust and selective quantification methods are required to better analyze feed supplementation effectiveness with specific amino acids. In this work, a reversed-phase high-performance liquid chromatography method with fluorescence detection is proposed and validated for lysine quantification, one of the most limiting amino acids in ruminant nutrition and essential towards milk production. To assess and widen method applicability, different matrices were considered: namely Li2CO3 buffer (the chosen standard reaction buffer), phosphate buffer solution (to mimic media in cellular studies), and rumen inoculum. The method was validated for all three matrices and found to be selective, accurate (92% ± 2%), and precise at both the inter- and intra-day levels in concentrations up to 225 µM, with detection and quantification limits lower than 1.24 and 4.14 µM, respectively. Sample stability was evaluated when stored at room temperature, 4 °C, and -20 °C, showing consistency for up to 48 h regardless of the matrix. Finally, the developed method was applied in the quantification of lysine on real samples. The results presented indicate that the proposed method can be applied towards free lysine quantification in ruminant feeding studies and potentially be of great benefit to dairy cow nutrition supplementation and optimization.

Dectin-1-Mediated Production of Pro-Inflammatory Cytokines Induced by Yeast ß-Glucans in Bovine Monocytes.

Yeast-derived products containing ß-glucans have long been used as feed supplements in domesticated animals in an attempt to increase immunity. ß-glucans are mainly recognized by the cell surface receptor CLEC7A, also designated Dectin-1. Although the immune mechanisms elicited through Dectin-1 activation have been studied in detail in mice and humans, they are poorly understood in other species. Here, we evaluated the response of bovine monocytes to soluble and particulate purified ß-glucans, and also to Zymosan. Our results show that particulate, but not soluble ß-glucans, can upregulate the surface expression of costimulatory molecules CD80 and CD86 on bovine monocytes. In addition, stimulated cells increased production of IL-8 and of TNF, IL1B, and IL6 mRNA expression, in a dose-dependent manner, which correlated positively with CLEC7A gene expression. Production of IL-8 and TNF expression decreased significantly after CLEC7A knockdown using two different pairs of siRNAs. Overall, we demonstrated here that bovine monocytes respond to particulate ß-glucans, through Dectin-1, by increasing the expression of pro-inflammatory cytokines. Our data support further studies in cattle on the induction of trained immunity using dietary ß-glucans.

The Use of Defatted Tenebrio molitor Larvae Meal as a Main Protein Source Is Supported in European Sea Bass (Dicentrarchus labrax) by Data on Growth Performance, Lipid Metabolism, and Flesh Quality.

OBJECTIVE: This study aims to determine the maximal inclusion level of defatted (d-) Tenebrio molitor larvae meal (TM) able to replace dietary fishmeal (FM) without compromising growth performance, general metabolism, and flesh quality traits in European sea bass, and to evaluate the major underlying physiological mechanisms. MATERIALS AND METHODS: Fish (55 ± 2 g) were fed with diets containing increasing levels of dTM: 0, 40, 80 and 100% (CTRL, TM40, TM80, and TM100, respectively) to replace FM. After 10 weeks of feeding, the growth performance, nutrient and energy balance, intestinal integrity, plasma metabolites and the expression of genes related to growth and nutrient metabolism, in liver and muscle were determined. The fatty acids (FA) profile, textural properties and color were also evaluated in muscle. RESULTS: Protein and lipids digestibility remained unaltered up to 80% dTM inclusion. Growth performance parameters were similar among dietary treatments. The dTM inclusion increased the hepatosomatic index in fish fed TM100. Muscle eicosapentaenoic acid, docosahexaenoic acid and n-3 long-chain polyunsaturated FA levels were maintained up to 80% dTM inclusion, but total cholesterol and non-esterified FA increased with dietary dTM inclusion. In liver, the expression of elongation of very long-chain FA protein 6 (elovl6) and FA desaturase 2 (fads2) did not change in fish fed TM40 and TM80, but elovl6 decreased whilst fads2 increased in fish fed TM100 when compared to those fed CTRL. The expression of cholesterol 7 alpha-monooxygenase (cyp7a1) decreased with dietary dTM inclusion. In muscle, the expression of myoblast determination protein-2 (myod2) decreased in fish fed TM80 and TM100. CONCLUSION: It is feasible to substitute dietary FM by dTM up to 80% in European sea bass without detrimental effects on nutrient digestibility, growth performance and associated genetic pathways, whilst assuring fillet nutritional value for human consumption.

Zinc in Dog Nutrition, Health and Disease: A Review.

Zinc is an essential trace element, required for enzymatic, structural, and regulatory functions. As body reserves are scarce, an adequate zinc status relies on proper dietary supply and efficient homeostasis. Several biomarkers have been proposed that enable the detection of poor zinc status, but more sensitive and specific ones are needed to detect marginal deficiencies. The zinc content of commercial dry dog foods has great variability, with a more frequent non-compliance with the maximum authorized limit than with the nutritional requirement. The bioavailability of dietary zinc also plays a crucial role in ensuring an adequate zinc status. Despite controversial results, organic zinc sources have been considered more bioavailable than inorganic sources, albeit the zinc source effect is more evident after a restriction period of dietary zinc. Many disorders have been associated with inadequate zinc status, not being clear whether the occurrence of the disease is the consequence or the cause. This review presents data on zinc requirements and biomarkers for zinc status, that can be applied for the development of supplementation strategies of zinc in complete pet foods. Moreover, it provides an understanding of the role zinc plays in the health of dogs, and how altered zinc status affects diseases in dogs.

Assessment of potato peel and agro-forestry biochars supplementation on in vitro ruminal fermentation.

BACKGROUND: The awareness of environmental and socio-economic impacts caused by greenhouse gas emissions from the livestock sector leverages the adoption of strategies to counteract it. Feed supplements can play an important role in the reduction of the main greenhouse gas produced by ruminants-methane (CH4). In this context, this study aims to assess the effect of two biochar sources and inclusion levels on rumen fermentation parameters in vitro. METHODS: Two sources of biochar (agro-forestry residues, AFB, and potato peel, PPB) were added at two levels (5 and 10%, dry matter (DM) basis) to two basal substrates (haylage and corn silage) and incubated 24-h with rumen inocula to assess the effects on CH4 production and main rumen fermentation parameters in vitro. RESULTS: AFB and PPB were obtained at different carbonization conditions resulting in different apparent surface areas, ash content, pH at the point of zero charge (pHpzc), and elemental analysis. Relative to control (0% biochar), biochar supplementation kept unaffected total gas production and yield (mL and mL/g DM, p = 0.140 and p = 0.240, respectively) and fermentation pH (p = 0.666), increased CH4production and yield (mL and mL/g DM, respectively, p = 0.001) and ammonia-N (NH3-N, p = 0.040), and decreased total volatile fatty acids (VFA) production (p < 0.001) and H2 generated and consumed (p ≤ 0.001). Biochar sources and inclusion levels had no negative effect on most of the fermentation parameters and efficiency. Acetic:propionic acid ratio (p = 0.048) and H2 consumed (p = 0.019) were lower with AFB inclusion when compared to PPB. Biochar inclusion at 10% reduced H2 consumed (p < 0.001) and tended to reduce total gas production (p = 0.055). Total VFA production (p = 0.019), acetic acid proportion (p = 0.011) and H2 generated (p = 0.048) were the lowest with AFB supplemented at 10%, no differences being observed among the other treatments. The basal substrate affected most fermentation parameters independently of biochar source and level used. DISCUSSION: Biochar supplementation increased NH3-N content, iso-butyric, iso-valeric and valeric acid proportions, and decreased VFA production suggesting a reduced energy supply for microbial growth, higher proteolysis and deamination of substrate N, and a decrease of NH3-N incorporation into microbial protein. No interaction was found between substrate and biochar source or level on any of the parameters measured. Although AFB and PPB had different textural and compositional characteristics, their effects on the rumen fermentation parameters were similar, the only observed effects being due to AFB included at 10%. Biochar supplementation promoted CH4 production regardless of the source and inclusion level, suggesting that there may be other effects beyond biomass and temperature of production of biochar, highlighting the need to consider other characteristics to better identify the mechanism by which biochar may influence CH4 production.