Diet supplementation with an organic acids-based formulation affects gut microbiota and expression of gut barrier genes in broilers

This study was designed to study the effect of diet supplementation with an organic acids-based formulation (OABF) on luminal- and mucosa-associated bacteria, concentration of volatile fatty acids (VFA), microbial glycolytic enzyme activity and expression of mucin 2 (MUC2), immunoglobulin A (IgA) and tight junction protein, i.e., zonula occludens-1 (ZO1), zonula occludens-2 (ZO2), claudin-1 (CLDN1), claudin-5 (CLDN5) and occludin (OCLN), genes at the ileal and cecal level. A 2 × 2 factorial design was used having OABF inclusion and avilamycin as main factors. Subsequently, 544 day-old male Cobb broilers were allocated in the following 4 treatments, each with 8 replicates: no additions (CON), 1 g OABF/kg diet (OA), 2.5 mg avilamycin/kg diet (AV) and combination of OA and AV (OAAV). The trial lasted for 42 days. In the ileum, OAAV resulted in lower mucosa-associated total bacteria levels (PO × A = 0.028) compared with AV. In addition, ileal digesta levels of Clostridium perfringens subgroup were decreased by avilamycin (PA = 0.045). Inclusion of OABF stimulated the activity of microbial glycolytic enzymes, whereas avilamycin resulted in lower acetate (PA = 0.021) and higher butyrate (PA = 0.010) molar ratios. Expression of ZO1 and CLDN5 was down-regulated by both OABF (PO = 0.016 and PO = 0.003, respectively) and avilamycin (PA = 0.016 and PA = 0.001, respectively). In addition, CLDN1 was down-regulated in AV compared with CON (PO × A = 0.012). Furthermore, OABF down-regulated MUC2 (PO = 0.027), whereas avilamycin down-regulated nuclear factor kappa B subunit 1 (NFKB1) (PA = 0.024), toll-like receptor 2 family member B (TLR2B) (PA = 0.011) and toll-like receptor 4 (TLR4) (PA = 0.014) expression. In the ceca, OABF inclusion increased digesta levels of Clostridium coccoides (PO = 0.018) and Clostridium leptum (PO = 0.040) subgroups, while it up-regulated MUC2 expression (PO = 0.014). Avilamycin (PA = 0.044) and interaction (PO × A < 0.001) effects for IgA expression were noted, with CON having higher IgA expression compared with AV. In conclusion, new findings regarding OABF inclusion effects on an array of relevant biomarkers for broiler gut ecology have been reported and discussed in parallel with avilamycin effects used as a positive control. This new knowledge is expected to provide a response baseline for follow up trials under various stress and challenge conditions. Keywords: Organic acids, Broiler chicken, Microflora, Tight junction, Microbial enzymes, Volatile fatty acid

Broiler gut microbiota and expressions of gut barrier genes affected by cereal type and phytogenic inclusion

The present study assessed the effects of cereal type and the inclusion level of a phytogenic feed additive (PFA) on broiler ileal and cecal gut microbiota composition, volatile fatty acids (VFA) and gene expression of toll like receptors (TLR), tight junction proteins, mucin 2 (MUC2) and secretory immunoglobulin A (sIgA). Depending on cereal type (i.e. maize or wheat) and PFA inclusion level (i.e. 0, 100 and 150 mg/kg diet), 450 one-day-old male broilers were allocated in 6 treatments according to a 2 × 3 factorial arrangement with 5 replicates of 15 broilers each, for 42 d. Significant interactions (P ≤ 0.05) between cereal type and PFA were shown for cecal digesta Bacteroides and Clostridium cluster XIVa, ileal digesta propionic and branched VFA, ileal sIgA gene expression, as well as cecal digesta branched and other VFA molar ratios. Cereal type affected the cecal microbiota composition. In particular, wheat-fed broilers had higher levels of mucosa-associated Lactobacillus (PCT = 0.007) and digesta Bifidobacterium (PCT < 0.001), as well as lower levels of total bacteria (PCT = 0.004) and Clostridia clusters I, IV and XIVa (PCT ≤ 0.05), compared with maize-fed ones. In addition, cereal type gave differences in fermentation intensity (PCT = 0.021) and in certain individual VFA molar ratios. Wheat-fed broilers had higher (P ≤ 0.05) ileal zonula occluden 2 (ZO-2) and lower ileal and cecal TLR2 and sIgA levels, compared with maize-fed broilers. On the other hand, PFA inclusion at 150 mg/kg had a stimulating effect on microbial fermentation at ileum and a retarding effect in ceca with additional variable VFA molar patterns. In addition, PFA inclusion at 100 mg/kg increased the ileal mucosa expression of claudin 5 (CLDN5) (PPFA = 0.023) and MUC2 (PPFA = 0.001) genes, and at 150 mg/kg decreased cecal TLR2 (PPFA = 0.022) gene expression compared with the un-supplemented controls. In conclusion, cereal type and PFA affected in combination and independently broiler gut microbiota composition and metabolic activity as well as the expression of critical gut barrier genes including TLR2. Further exploitation of these properties in cases of stressor challenges is warranted. Keywords: Maize, Wheat, Phytogenics, Gut microbiota, Toll like receptors, Gut barrie

Phytogenic Administration and Reduction of Dietary Energy and Protein Levels Affects Growth Performance, Nutrient Digestibility and Antioxidant Status of Broilers

The aim of this study was to investigate the effect of reduced dietary energy (ME) and protein (CP) levels along with administration of a phytogenic feed additive (PFA) based on oregano, anise and citrus essential oils, on broiler growth performance, nutrient digestibility, meat and blood biochemical parameters and total antioxidant capacity (TAC). Depending on dietary ME and CP level down regulation compared to a corn-soybean meal basal diet A used as positive control, three diet types [A, B(＝A-3%) and C(＝A-6%)] were implemented. Depending on the inclusion or not of PFA at 125 mg/kg diet, 450 1-d old, male Cobb broilers were randomly allocated in six treatments according to a 3×2 factorial arrangement with 5 replicates of 15 broilers; A: diets formulated optimally to meet broiler nutrient requirements for maximizing protein content of meat for starter, grower and finisher growth periods; APh: A+PFA; B: suboptimal in ME and CP levels by 3%; BPh: B+PFA; C: suboptimal in ME and CP levels by 6%; CPh: C+PFA. Feed conversion ratio (FCR) was improved in birds fed diet A compared to diet C during the grower period (PD＝0.021) and overall (PD＝0.010). Phytogenic supplementation resulted in higher (PD×Ph＝0.020) total tract apparent digestibility of fat in birds fed diet C compared to diet A. Birds fed diet A had higher (PD＝0.001) plasma cholesterol, compared to birds fed diet C. In addition, birds fed diets A and B had higher (PD＝0.002) breast protein content compared to C. Overall, PFA inclusion reduced cholesterol (PPh＝0.002) and increased plasma TAC (PPh&lt;0.001). Moreover, PFA increased breast (PPh＝0.001) and thigh (PPh＝0.01) TAC. In conclusion, a reduction in dietary ME and CP levels, adversely affected the FCR, whereas PFA supplementation tended (PPh＝0.089) to compensate these effects. Moreover, the addition of PFA reduced plasma cholesterol and improved plasma and meat TAC

Growth performance, nutrient digestibility, antioxidant capacity, blood biochemical biomarkers and cytokines expression in broiler chickens fed different phytogenic levels

The effects of inclusion levels of a phytogenic feed additive (PFA), characterized by menthol anethol and eugenol, on broiler growth performance, nutrient digestibility, biochemical biomarkers and total antioxidant capacity (TAC) of plasma and meat, as well as on the relative expression of selected cytokines, were studied in a 42-d experiment. A total of 225 one-day-old male Cobb broiler chickens were assigned into 3 treatments, with 5 replicates of 15 chickens each. Chickens were fed maize-soybean meal basal diets following a 3 phase (i.e., starter, grower and finisher) feeding program. Depending on PFA inclusion level, treatments were: no PFA (PFA-0), PFA at 100 mg/kg (PFA-100) and PFA at 150 mg/kg (PFA-150). Feed and water were available ad libitum. Feed conversion ratio (FCR) during finisher phase was improved quadratically (P < 0.05) with increasing PFA level. Overall, increasing PFA level increased body weight gain (BWG) in a linear (P < 0.05) and quadratic (P < 0.05) manner with treatments PFA-100 and PFA-150 being greater (P < 0.05) compared with PFA-0. Total tract apparent digestibility of dry matter increased linearly (P < 0.05) and quadratically (P < 0.05) with increasing PFA level. The apparent metabolizable energy corrected for nitrogen (AMEn) also increased linearly (P < 0.05). Increasing PFA level resulted in a linear (P < 0.05) increase in blood plasma TAC. Expression of pro-inflammatory cytokine interleukin -18 (IL-18) was reduced linearly (P < 0.05) in spleen with increasing PFA level. In conclusion, PFA inclusion at 100 mg/kg diet positively influenced performance, whereas PFA inclusion at 150 mg/kg resulted in a stronger improvement in AMEn and plasma TAC. Finally, PFA inclusion resulted in a pattern of reduced pro-inflammatory biomarker IL-18 at spleen. Overall, this study provides evidence for the beneficial role of PFA as a natural growth and health promoter in broiler chickens that needs to be further confirmed in field studies

Effects of deoxynivalenol and fumonisins on broiler gut cytoprotective capacity

Mycotoxins are a crucial problem for poultry production worldwide. Two of the most frequently found mycotoxins in feedstuffs are deoxynivalenol (DON) and fumonisins (FUM) which adversely affect gut health and poultry performance. The current knowledge on DON and FUM effects on broiler responses relevant for gut detoxification, antioxidant capacity, and health is still unclear. The aim of this study was to assess a range of selected molecular intestinal biomarkers for their responsiveness to the maximum allowable European Union dietary levels for DON (5 mg/kg) and FUM (20 mg/kg) in broilers. For the experimental purpose, a challenge diet was formulated, and biomarkers relevant for detoxification, antioxidant response, stress, inflammation, and integrity were profiled across the broiler intestine. The results reveal that DON significantly (p < 0.05) induced aryl hydrocarbon receptor (AhR) and cytochrome P450 enzyme (CYP) expression mainly at the duodenum. Moreover, DON and FUM had specific significant (p < 0.05) effects on the antioxidant response, stress, inflammation, and integrity depending on the intestinal segment. Consequently, broiler molecular responses to DON and FUM assessed via a powerful palette of biomarkers were shown to be mycotoxin and intestinal site specific. The study findings could be highly relevant for assessing various dietary bioactive components for protection against mycotoxins

Evaluation of a Natural Phytogenic Formulation as an Alternative to Pharmaceutical Zinc Oxide in the Diet of Weaned Piglets

A natural phytogenic formulation (NPF) was tested as an alternative to pharmaceutical zinc oxide (ZnO) in weaned piglets with respect to growth performance, apparent total tract digestibility and faecal microbiota composition and metabolic activity. Two dietary NPF levels (NPF: 1000 and 2000 mg/kg diet) were compared to a positive control (ZnO: 3000 mg ZnO/kg diet) and a negative control (CON: no added ZnO or NPF) using 84 weaned piglets from 29 d to 78 d (days of age). Feed conversion ratio was improved (p &lt; 0.05) in ZnO and NPF piglets were compared to CON at 50 d. Dry matter, organic matter and crude protein (p &lt; 0.05) digestibility was improved in NPF piglets compared to CON at 57 d. Compared to CON, NPF inclusion reduced E. coli (p &lt; 0.05) and increased C. leptum subgroup (p &lt; 0.01) at 57 d and 78 d, and reduced C. perfringens subgroup (p &lt; 0.05; at 78 d). The ZnO reduced (p &lt; 0.001) E. coli and C. perfringens subgroup (p &lt; 0.01) compared to CON at 78 d. Moreover, ZnO and NPF reduced molar ratios of branched chain volatile fatty acids (p &lt; 0.05) compared to CON, while NPF also increased butyric acid (p &lt; 0.05) at 78 d. In conclusion, the NPF appeared to be a promising alternative to pharmaceutical doses of ZnO

Phytogenic Effects on Layer Production Performance and Cytoprotective Response in the Duodenum

The aim of this study was to evaluate the effects of a phytogenic premix (PP) on the production performance and critical genes relevant to the detoxification (i.e., aryl hydrocarbon receptor pathway) and antioxidant (i.e., nuclear factor erythroid 2-related factor 2 pathway) response in the duodenum of laying hens. The PP was based on bioactive substances derived from ginger, lemon balm, oregano, and thyme (Anco FIT-Poultry). A total of 385 20 week old Hy-Line Brown layers were assigned to five dietary treatments with seven replicates of 11 hens each for a 12-week feeding trial. The experimental treatments included a corn–soybean meal basal diet with no PP (CON) or supplemented with PP at 500 (P500), 750 (P750), 1000 (P1000), and 1500 mg/kg diet (P1500). The overall (1–12 weeks) laying rate (p p = 0.008) were significantly increased in the P1000 group compared with the CON. At the duodenum, increasing dietary PP inclusion levels beneficially affected (p ≤ 0.05) the expression of the majority of the AhR and Nrf2 pathway genes studied. In conclusion, according to the gene expression analysis, PP inclusion resulted in a reduced requirement for detoxification and an increased antioxidant capacity, with most of the effects seen at the PP inclusion range of 750 to 1000 mg/kg diet

Intestinal Bacteria Composition and Translocation of Bacteria in Inflammatory Bowel Disease.

Live commensal intestinal bacteria are present in the peripheral blood where they can induce inflammation.To evaluate the intestinal bacteria composition and translocation of bacteria in IBD.Both blood and tissue biopsy samples were collected from adult patients with active/inactive Crohn's disease (CD), active/inactive ulcerative colitis (UC) and healthy individuals. Most of the patients were newly diagnosed and none of them received antibiotics. Using a reverse transcription-quantitative real-time PCR (RT-qPCR) method, we determined the composition of microbiota. NOD2/CARD15 genotyping was also studied.Total bacterial DNA concentration was increased in tissue and blood samples of IBD patients compared to healthy controls. Furthermore, the active IBD cases had higher total bacterial DNA concentration levels compared to the inactive cases. Three species characterized dysbiosis in IBD, namely an increase of Bacteroides spp in active and inactive IBD samples, and a decrease in Clostridium leptum group (IV), and Faecalibacterium prausnitzi in both active and inactive IBD patients. No significant association between bacterial translocation and NOD2/CARD15 mutations was found.The composition of the microbiota in IBD patients differs from that of healthy controls. The high rate of bacterial DNA in the blood samples indicates translocation in inflammatory bowel disease