Relative bioefficacy of RRR-α-tocopherol versus all-rac-α-tocopherol in in vitro models

The aim of this study was to evaluate the in vitro relative bioefficacy of RRR-α-tocopherol (RRR- α-T) versus all-rac-α-tocopherol (all-rac-α-T) in counteracting the cytotoxic effect induced by H2O2 in Bovine Mammary Epithelium – University of Vermont (BME-UV1) and Madin-Darby Canine Kidney (MDCK) cells. The range of RRR- α-T and all-rac- α-T concentrations selected for the oxidative challenge experiments was 100µM - 1nM. To study the bioefficacy of RRR- α-T and all-rac- α-T, MTT and LDH tests were performed. Cells were pre-incubated for 3 h with  selected a-tocopherol concentrations and then exposed to increasing H2O2 concentrations ranging from 125 to 750µM for the following 24h. Concerning the cell viability, the pre-treatments with 100µM of RRR- α-T and 100µM all-rac-α-T were able to significantly (P<0.05) counteract the effect induced by 750 µM of H2O2 in BME-UV1. In MDCK the pre-treatment with 1nM of all-rac-α-T was able to significantly (P<0.05) reduce the effect of 125 and 150 mM H2O2. In MDCK cells, the pre-incubation with all-rac-α-T determines a significant reduction of the membrane damage, induced by 175 µM of H2O2. In conclusion, RRR-α-T and all-rac-α-T have shown the ability to counteract the oxidative effects of H2O2, however further investigation will help to better understand their specific mechanism of action in vitro.  

Effect of Zinc Oxide and Zinc Chloride on Human and Swine Intestinal Epithelial Cell Lines

Zinc (Zn) salts are often used as nutritional additives in order to promote gut health. The aim of the present study was to assess the effect of two widely used additives in feedstuff, on the intestinal epithelium. In particular, the effect of zinc oxide (ZnO) and zinc chloride (ZnCl2) was investigated in human (INT-407) and porcine (IPI-2I) cell line models. The effect of Zn sources on IPI-21 and INT-407 cell lines was evaluated by a colorimetric viability test using an incubation period of 3 and 24 hours under serum-free conditions. INT407 and IPI-2I showed to be a suitable model of the intestine and a simple tool to investigate the role of Zn supplements. INT407 showed to be the most sensible model to Zn supplements considered, whereas IPI-2I were more resistant. The results of this study contribute to determine the role of zinc in human and swine intestinal epithelium. However, further in vivo experiments may be done to clarify the contribution of Zn supplements in gut health and to improve Zn supplementation in animal feed and in human formulations

In vitro evaluation of antimicrobial and antioxidant activities of algal extracts

The aim of this study was to evaluate the antioxidant capacity, the antimicrobial proprieties of algae Ascophyllum nodosum and Schizochytrium spp. against one of major swine enteric pathogen Escherichia coli O138 by broth macro-dilution method in Luria-Bertani (LB) medium. The antimicrobial effect of the algal extracts at supplementation of 0.12%, 0.06% and 0.03% (v/v) on E. coli O138, genetically characterised by PCR, was evaluated by following the bacterial growth. The antioxidant activity was determined by the ABTS Radical Cation Decolorisation Assay. In particular, the log(10) E. coli used as control resulted significantly higher than 0.12% at 3 hours (8.82 +/- 0.07 and 8.18 +/- 0.07 log(10) cells/mL, respectively; p<.01) suggesting an inhibitory activity related to the dose. No effect activity was observed with Schizochytrium spp. against E. coli growth. A. nodosum and Schizochytrium spp. exhibited antioxidant capacity (p<.05). The combination of them (1:1) exhibited antioxidant activity suggesting a synergistic effect (p<.05). The different proprieties of algal species that can modulate the O138 E. coli growth, one of the major pathogen of swine species, together with the antioxidant capacity, make them a promising functional feed additive to improve the gut health, therefore further studies are needed to confirm these activities in vivo

Role of omega-3 polyunsaturated fatty acids, citrus pectin, and milk-derived exosomes on intestinal barrier integrity and immunity in animals

The gastrointestinal tract of livestock and poultry is prone to challenge by feedborne antigens, pathogens, and other stress factors in the farm environment. Excessive physiological inflammation and oxidative stress that arises firstly disrupts the intestinal epithelial barrier followed by other components of the gastrointestinal tract. In the present review, the interrelationship between intestinal barrier inflammation and oxidative stress that contributes to the pathogenesis of inflammatory bowel disease was described. Further, the role of naturally existing immunomodulatory nutrients such as the omega-3 polyunsaturated fatty acids, citrus pectin, and milk-derived exosomes in preventing intestinal barrier inflammation was discussed. Based on the existing evidence, the possible molecular mechanism of these bioactive nutrients in the intestinal barrier was outlined for application in animal diets

Administration of potentiated Zn and monovalent Cu in weanling piglets diet

The aim of this study was to determine the suitability of potentiated Zn combined with a monovalent Cu as a replacement for pharmacological ZnO supplementation in piglets after weaning. A total of 120 piglets (7.143 ± 0.924 kg) were weaned at 27d and divided into 4 experimental treatments. A positive control (PC, 2500 ppm of Zinc through standard ZnO) was compared to 3 treatments in which Cu and Zn were supplemented through potentiated Zn (HiZox®) and monovalent Cu (CoRouge®) at European and non-European levels of inclusion: EU (120 ppm of Zn; 140 ppm of Cu), non-EU+ (300 ppm of Zn; 200 ppm of Cu) and non-EU- (300 ppm of Zn; 140 ppm of Cu). Performance data were analyzed using the MIXED procedure of SAS. Serum diamine oxidase (DAO) and D-lactate were analysed through a MIXED procedure of SAS at 0d and 14d. SIgA was evaluated with a GLM procedure of SAS (28d). Fecal samples (14d and 28d) and intestinal content (28d) were collected and the V3–V4 hypervariable regions of the bacterial 16S gene were sequenced in one MiSeq (Illumina) run. Performances were not affected by different Zn and Cu dosages. Plasma DAO was negatively (p<0.05) affected in non-EU- treatment. The level of sIgA was increased in PC group (157.73 ± 19.58 ng/ml) compared to EU (143.81 ± 12.96 ng/ml; p<0.05) and non-EU- group (138.01 ± 16.88 ng/ml; p<0.01). non-EU- contributed to a significant decrease in biodiversity in fecal microbiota as shown by the Shannon diversity and Simpson evenness indexes (p<0.05). A total of 18 differentially abundant genera (p<0.05) were identified in the caecal content with the presence of genera linked to the disruption of the gut barrier (Campylobacter) with non-EU- . These results suggest that a more balanced supplementation of the two trace elements, through more bioavailable sources, could represent a valid tool to enhance gut health of weaning piglets and reduce environmental impact

The relationship between potentiated zinc oxide and monovalent copper in a dietary administration for weanling piglets

The aim of this study was to evaluate different dietary level of Zinc (Zn) and Copper (Cu) supplementation according to European and non-European levels on the health and performance of pigs 14 d after weaning. A total of 120 piglets [initial body weight = 7.14 ± 0.92 kg] were divided into 4 experimental treatments: positive control (PC, 2,500 ppm of Zn, ZnO) and 3 treatments in which Zn and Cu were added through potentiated ZnO and Cu2O according to different European and non-European levels of inclusion: EU (120 ppm of Zn; 140 ppm of Cu), non-EU+ (300 ppm of Zn; 200 ppm of Cu) and non-EU- (300 ppm of Zn; 140 ppm of Cu). Growth performance, serum Zn and Cu concentrations, fecal score, blood biomarkers of intestinal integrity, and fecal microbial composition were examined and analyzed using GLM and MIXED procedures of SAS. Fecal samples at 14 d were collected and the V3–V4 hypervariable regions of the bacterial 16S gene were sequenced in one Illumina MiSeq run. Body weight (BW), average daily gain (ADG), average daily feed intake (ADFI), feed conversion ratio (FCR) and (feed efficiency (FE) were not affected by the experimental treatments. The fecal score suggests that during the first 14d, the consistency of the feces tended (P < 0.10) to be less in the three treatment groups while in PC, the greatest concentration of Zn favored a better fecal consistency. The initial serum Zn and Cu concentrations were similar for all dietary treatment groups. After 14 d, serum Zn concentration was significantly greater in PC (15.71 ± 6.25 μg/L, P < 0.05). No differences were highlighted in serum Zn concentration among the other 3 treatment groups. Serum Cu concentrations revealed statistically significant results (P < 0.05) on 14 d after weaning in EU, non-EU+ and non-EU- groups, showing greater concentrations of Cu (17.65 ± 5.49 μmol/L, 17.73± 3.46 μmol/L 18.33 ± 4.13 μmol/L, respectively) compared with PC (11.87 ± 3.60 μmol/L). These results suggested that the EU level of inclusion of both potentiated Zn and monovalent Cu is enough to reach the same Cu and Zn plasma concentrations of non-EU treatments. Plasma DAO was negatively (P < 0.05) affected in non-EU-. Firmicutes and Bacteroidetes were the most abundant phyla in fecal microbiome. Non-EU- contributed to a significant decrease in biodiversity in fecal microbiota as depicted with Shannon for diversity and Simpson for evenness indexes (P < 0.05). In fecal samples, the presence of genera linked to a greater disruption of the gut barrier (Escherichia-Shigella) was depicted in non-EU- group, indicating significant modifications of the microbial community. These results suggest the need for a balanced supplementation of Cu and Zn through more bioavailable sources. The use of potentiated ZnO and Cu2O according to European levels could represent a valid strategy to enhance weanling piglets gut health and to reduce the environmental impact

A herbal vitamin D source improves the performance of piglets fed low-calcium starter diets

Weaning is a stressful period for piglets that can negatively affect their performance. Plants contain a variety of secondary metabolites that act as antioxidants or anti-inflammatory agents that can support piglets after weaning. In a trial with 192 crossbred Topigs weaned at 27 days of age, effects of Solanum glaucophyllum (SG) alone or in combination with a mixture of essential oils (EO) on the performance of weaning pigs fed calcium (Ca)-reduced diets was tested. The piglets were kept in 48 pens and fed one of four diets for 35 days, with a prestarter diet from d0-14 and a starter diet from d14-35: control (PC), a standard diet; negative control (NC), diet with reduced Ca and phosphorus; NC supplemented with SG (SG) and NC supplemented with SG and EO (SG+). Individual and pen body weight were measured on d0, d14, and d35, and pen feed intake was measured on d14 and d35. Body weight and daily weight gain did not differ among the treatments. In the prestarter period, SG had the lowest FCR (1,539 ± 0,127; p < 0.05). In the starter period, the FCR of NC (1.455 ± 0.144) was significantly (p < 0.05) higher than that of SG+ (1.297 ± 0.195), while the FCR of the two other diets was intermediate. The results indicate a better feed use in diets with lower dietary Ca. Adding SG alone had a strong effect on post-weaning performance, while combining SG and EO showed minor synergies. Additional analysis to better understand the underlying mechanisms is currently underway

Functional characterisation of Euglena gracilis following growth medium enrichment

In recent years, microalgae, particularly Euglena gracilis, have been candidates for the food/feed industry thanks to their nutritional and functional properties. However, the inclusion of E. gracilis in the diet of livestock animals is little studied, especially regarding its antioxidant activity. Furthermore, microalgae are known for their variability in nutritional quality and functional properties, mainly due to cultivation conditions. For this reason, the aim of the present work was to investigate the nutritional and functional aspects (total phenolic content TPC, Folin-Ciocalteu assay); antioxidant activity (ABTS and FRAP assays) of E. gracilis, grown in three different media, following chemical extraction (H2O:EtOH) and ex vivo digestion method. The microalgae growth media were characterised as follows: EgM (standard medium); ETX (standard medium + aminoacidic-extract); DOE-ETX (ETX + microelements). The results showed an interesting nutritional profile for all the microalgae analysed, although the values were modulated by media nutrients. Although EgM (6.94 +/- 0.25 g/L) was characterised by a significant higher growth yield than ETX (5.75 +/- 0.14 g/L) and DOE-ETX (4.72 +/- 0.17 g/L), results also confirmed by the paramylon content (4.35 +/- 0.13; 3.16 +/- 0.08; 2.25 +/- 0.05 g/L, respectively) (p < 0.05), it did not show a high functional profile. More specifically, DOE-ETX showed higher values for TPC, ABTS, and FRAP, following chemical extraction, in particular 50 and 75% EtOH and ex vivo digestion. These results confirmed the potential of E. gracilis as a valuable source of functional feed ingredients. Further investigations will be crucial to optimise the formulation of the culture medium to obtain a high yield of algae with improved functional characteristics

Evaluation of antigens stability of tobacco seeds as edible vaccine against VTEC strains

Plants have represent a promising alternative for biopharmaceutical proteins (Ma et al., 2003; Rossi et al., 2014). Many plant based edible vaccines have been shown to be effective in inducing local immune responses (Rossi et al., 2013). Edible vaccines can activate both mucosal and systemic immunity, as they come in contact with the digestive tract lining. This dual effect would provide first-line defense against pathogens invading through the mucosa. The antigens are released in the intestines are taken up by M cells that are present over the Payer’s patches (in the ileum) and the gut associated lymphoid tissue (GALT). Edible vaccines represent an important worldwide goal for the prevention of the enteric diseases, also in livestock. In particular, the enteric infections are a significant clinical problem in pigs. Verocytotoxic Escherichia (E.) coli strains are responsible for serious enterotoxaemia that causes important economic losses in the pig industry. The production of a vaccine for oral administration of transgenic seeds could be a practical and efficient system to prevent the infection and to reduce the antibiotic use. This study was focused on tobacco plants, previously transformed by agroinfection for the seed-specific expression of antigenic proteins (F18 adhesive fimbriae and the B subunit of the Vt2e toxin) as model of edible vaccines against verocytotoxic E. coli strains. The dietary administration of transgenic tobacco seeds promotes a significant increase in the number of mucosal IgA-producing cells of the tunica propria in both small and large intestine in mice (Rossi et al., 2013). A protective effect of oral administration of transgenic tobacco seeds was also observed against verocytotoxic Escherichia coli infection in piglets (Rossi et al., 2014). The aim of this study was to assess the seed-expression stability, that is a important requirement in the vaccine production, of F 18 and Vt2e-B heterologous genes into the progeny of transformed tobacco plants