Effects of a dietary isoquinoline alkaloids blend on gut antioxidant capacity and gut barrier of young broilers.

Extensive mechanistic evidence to support the beneficial function of dietary phytobiotic applications for broiler performance, gut function and health is highly warranted. In particular, for isoquinoline alkaloids (IQ) the underlying mechanisms related to critical gut homeostasis components such as cytoprotection and gut barrier are scarce, especially for young broilers at the starter growth stage (d1-10). The aim of this study was to investigate the effect of a standardized blend of IQs on the relative gene expression of critical biomarkers relevant for antioxidant response and barrier function along the intestine of young broilers at the end of starter growth phase. For this purpose, 182 one-day-old Ross 308 broilers were allocated in 2 treatments with 7 replicates of 13 broilers each: control diet-no other additions (NC), and control diet containing a standardized blend of IQs at 200 mg/kg of diet (M) for the starter growth period (1-10d). The results revealed that the IQs blend significantly upregulated (P < 0.05) the expression of genes related to antioxidant response in all intestinal segments. Moreover, the IQs blend enhanced (P < 0.05) gut barrier components primarily at duodenal level. In conclusion, the blend of IQs beneficially affected critical pathway components relevant for the gut antioxidant capacity and barrier along the intestine of young broilers.

Dietary Inclusion Level Effects of Yoghurt Acid Whey Powder on Performance, Digestibility of Nutrients and Meat Quality of Broilers.

In recent years, the increasing demand for Greek strained yoghurt produced massive amounts of acid whey, which constitutes a major environmental pollutant. Whether yoghurt acid whey can be included in poultry diets is not known. The purpose of this study was to evaluate the effects of four dietary levels of yoghurt acid whey powder (YAWP) on the growth performance, nutrient digestibility, meat quality traits and oxidative stability. A total of 300 male 1-day-old Ross 308 broilers were assigned into four groups with five replicates of 15 broilers each: control-fed basal diet with no YAWP addition (WO) or basal diet supplemented with YAWP at 25 g/kg of diet (WA), 50 g/kg of diet (WB), or 100 g/kg of diet (WC). At the starter period, body weight and body weight gain were reduced after WB and WC treatments compared to the WO treatment. Breast meat oxidative stability was improved during refrigerated storage for 1 and 3 d in all YAWP treatments compared to control, while the WA treatment showed an improved oxidative stability after 6 and 9 d. The results suggest that YAWP inclusion at 25 g/kg of diet did not impair performance and extended the meat shelf life by reducing lipid oxidation rates.

Egg White and Yolk Protein Atlas: New Protein Insights of a Global Landmark Food.

(1) Background: The chicken egg is an animal product of great agronomic interest. The egg white and yolk constitute high-quality protein sources for humans with high digestibility and well-balanced amino acid profiles. Despite the egg white and yolk protein's undisputed value, research to unravel their full proteome content and its properties is still ongoing. We aimed to exhaustively analyze the proteome of egg white and yolk by applying intrinsic proteomics and bioinformatics approaches in order to unravel the full protein potential of this landmark food. (2) Methods: A total of 45 freshly laid, unfertilized, chicken eggs were subjected to nanoLC-MS/MS Orbitrap analysis following a peptide pre-fractionation step. A comprehensive bioinformatics processing step was undertaken towards elucidating potential activities and roles of identified molecules. In parallel, the literature was mined concerning all reported egg white and yolk protein identifications. (3) Results: Our analysis revealed 371 and 428 new proteins, reported for the first time to be present in the egg white and yolk, respectively. From the bioactivity standpoint, egg white and yolk proteins showed high enrichment for antioxidant and anti-inflammatory processes, while exerting high relevance for the apoptosis and focal adhesion pathways. (4) Conclusions: Egg white and yolk proteins exert diverse and multifaceted properties. A total of 799 proteins were reported for the first time as being part of the egg and yolk. Our novel protein data enriched those already published in the literature and the first ever chicken egg white and yolk Protein Atlas, comprising 1392 protein entries, was generated. This dataset will provide a cornerstone reference for future studies involving egg proteins.

Nutrigenomic profiling of reduced specification diets and phytogenic inclusion effects on critical toll-like receptor signaling, mitogen-activated protein kinase-apoptosis, and PI3K-Akt-mTOR gene components along the broiler gut.

The effects of concurrent reduction of dietary metabolizable energy (ME) and crude protein (CP) levels combined or not with the dietary inclusion of a phytogenic feed additive (PFA) were studied using a nutrigenomics approach. In particular, the expression of 26 critical genes relevant for inflammation control (TLR pathway), cellular apoptosis (MAPK pathway) cell growth and nutrient metabolism (PI3K-Akt-mTOR pathway) was profiled along the broiler intestine. Two dietary types (L and H) differing in metabolizable energy and crude protein levels (L: 95% and H: 100% of optimal Cobb 500 recommendations for ME and CP requirements) supplemented or not with PFA (- or +) and their interactions (L-, L+, H-, H+) were evaluated. There were only 3 total interactions (mTOR, IL8, and HRAS P < 0.05) between diet type and PFA inclusion indicating limited concurrent effects. Diet type, L upregulated genes related with inflammation mainly in the jejunum, ileum, and cecum (P < 0.05) and MAPK pathway in the ileum and cecum (P < 0.05). Moreover, diet type L negatively affected the expression of genes related to PI3K-Akt-mTOR pathway mainly in duodenum and cecum (P < 0.05). On the other hand, PFA inclusion downregulated (P < 0.05) genes related with TLR signaling pathway (TLR2B, MyD88, TLR3, IL8, LITAF) along the intestine and MAPK pathway genes (APO1, FOS) in jejunum (P < 0.05). Finally, PFA supplementation regulated nutrient sensing and metabolism in the cecum in a manner perceived as beneficial for growth. In conclusion, the study results highlight that the reduced ME and CP specifications, especially in the absence of PFA, regulate inflammation, apoptosis and nutrient metabolism processes at homeostatic control levels that hinder maximizing the availability of dietary energy and nutrients for growth purposes. Inclusion of PFA helped to adjust the respective homeostatic responses and control to levels supporting broiler performance, especially at reduced specification diets.

In Vitro Fermentation of Edible Mushrooms: Effects on Faecal Microbiota Characteristics of Autistic and Neurotypical Children.

Children with autism spectrum disorder (ASD) often suffer gastrointestinal disturbances consistent with gut microbiota (GM) alterations. Treatment with pro/prebiotics may potentially alleviate gut symptoms, but the evidence for prebiotics is scarce. This study aims to evaluate the effects of edible mushrooms (Pleurotus, Basidiomycota) and prebiotic compounds on GM composition and metabolite production in vitro, using faecal samples from autistic and non-autistic children. Specific microbial populations were enumerated after 24 h of fermentation by quantitative PCR, and the metabolic production was determined by gas chromatography. Higher levels of Prevotella spp. and Bifidobacterium spp. were measured in neurotypical children compared to ASD children. A total of 24 h fermentation of Pleurotus eryngii and P. ostreatus mushroom powder increased the levels of Bifidobacterium, while known prebiotics increased the levels of total bacteria and Bacteroides in both groups. Only P. eryngii mushrooms resulted in significantly elevated levels of total bacteria Bacteroides and Feacalibacterium prausnitzii compared to the negative control (NC) in the ASD group. Both mushrooms induced elevated levels of butyrate after 24 h of fermentation, while short-chain fructooligosaccharides induced increased levels of acetate in the ASD group, compared to NC. Overall, this study highlights the positive effect of edible mushrooms on the GM and metabolic activity of children with ASD.

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 < 0.05) in ZnO and NPF piglets were compared to CON at 50 d. Dry matter, organic matter and crude protein (p < 0.05) digestibility was improved in NPF piglets compared to CON at 57 d. Compared to CON, NPF inclusion reduced E. coli (p < 0.05) and increased C. leptum subgroup (p < 0.01) at 57 d and 78 d, and reduced C. perfringens subgroup (p < 0.05; at 78 d). The ZnO reduced (p < 0.001) E. coli and C. perfringens subgroup (p < 0.01) compared to CON at 78 d. Moreover, ZnO and NPF reduced molar ratios of branched chain volatile fatty acids (p < 0.05) compared to CON, while NPF also increased butyric acid (p < 0.05) at 78 d. In conclusion, the NPF appeared to be a promising alternative to pharmaceutical doses of ZnO.

Dietary phytogenic inclusion level affects production performance and expression of ovarian cytoprotective genes in laying hens.

A 12-wk study was conducted to investigate the effects of a phytogenic premix (PP) inclusion level on production performance, and the expression of genes relevant for detoxification (aryl hydrocarbon receptor; AhR) and antioxidant capacity (Nuclear factor erythroid 2-related factor 2; Nrf2) in the ovaries of laying hens. The PP consisted of bioactive substances derived from ginger, lemon balm, oregano, and thyme substances (Anco FIT-Poultry). Depending on PP inclusion level (i.e., 0, 500, 750, 1,000, and 1,500 mg/kg diet) in the basal diet, 385 laying hens Hy-Line Brown, 20-wk-old were assigned into 5 treatments: CON, P500, P750, P1000, and P1500, with 7 replicates of 11 hens with ad libitum access to feed and water. Performance parameters were closely monitored on a weekly basis and analyzed in the following 3 experimental periods: 1 to 4 wk, 5 to 8 wk, and 9 to 12 wk of treatment administration (i.e., 21-24, 25-28, and 29-32 wk of layers age, respectively). At the end of the 8th and 12th wk of the experiment (i.e., 28 and 32 wk of layers age), a layer from each replicate was selected, euthanized, the ovaries sampled and stored deep frozen until gene expression analysis. Data were analyzed by ANOVA and means compared using Tukey's honest significant difference test. Polynomial contrasts tested the linear and quadratic effect of PP inclusion levels. Results revealed that PP inclusion, improved (P < 0.05) laying rate and egg mass, compared to CON. Increasing PP inclusion level enhanced laying rate and egg mass, linearly and quadratically and peaked at P1000 (P < 0.05). In the ovaries, the AhR pathway genes assessed were down-regulated (P < 0.05) mainly at P1000 and P750 treatments. In addition, PP related cytoprotective potential was demonstrated via beneficial changes seen for the majority of the Nrf2-pathway genes assessed with the P1000 displaying most significant differences from CON. Conclusively, new data highlighted beneficial cytoprotective effects of PP inclusion on layer ovaries and documented further layer performance, with the inclusion level of 1000 mg PP/kg diet being the most prominent.

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 < 0.001) and egg mass (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.

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.

Effects of Diet and Phytogenic Inclusion on the Antioxidant Capacity of the Broiler Chicken Gut.

The reduction in energy and protein dietary levels, whilst preserving the gut health of broilers, is warranted in modern poultry production. Phytogenic feed additives (PFAs) are purported to enhance performance and antioxidant capacity in broilers. However, few studies have assessed PFA effects on a molecular level related to antioxidant response. The aim of this study was to investigate the effects of administering two dietary types differing in energy and protein levels (L: 95% and H: 100% of hybrid optimal recommendations) supplemented with or without PFA (-, +) on gene expressions relevant for antioxidant response along the broiler gut. Interactions of diet type with PFA (i.e., treatments L-, L+, H-, H+) were determined for critical antioxidant and cyto-protective genes (i.e., nuclear factor erythroid 2-like 2 (Nrf2) pathway) and for the total antioxidant capacity (TAC) in the proximal gut. In particular, the overall antioxidant response along the broiler gut was increased upon reduced dietary energy and protein intake (diet type L) and consistently up-regulated by PFA addition. The study results provide a new mechanistic insight of diet and PFA functions with respect to the overall broiler gut antioxidant capacity.

Fermentation of Pleurotus ostreatus and Ganoderma lucidum mushrooms and their extracts by the gut microbiota of healthy and osteopenic women: potential prebiotic effect and impact of mushroom fermentation products on human osteoblasts.

Recent data have highlighted the role of the gut microbiota and its several metabolites in maintaining bone health. Thus, gut microbiota manipulation, e.g., by prebiotics, might offer a plausible target in the fight against bone degenerative diseases. This study aimed (a) to investigate the in vitro prebiotic potential of Ganoderma lucidum and Pleurotus ostreatus mushrooms in healthy and osteopenic women and (b) to explore the impact of mushroom fermentation products on human osteoblasts. G. lucidum LGAM 9720 and P. ostreatus IK 1123 lyophilized mushroom-powders (2% w/v) and their hot-water extracts (1% w/v) were fermented in a 24 h static batch culture model by using faecal inocula from healthy (n = 3) or osteopenic (n = 3) donors. Gut microbiota analysis (qPCR) and measurement of short chain fatty acids (SCFAs) were performed during fermentation, and 24 h-prebiotic indexes were calculated. Evaluation of the effects of fermentation products on bone metabolism parameters (OPG: osteoprotegerin; and RANKL: receptor activator of nuclear factor kappa B ligand) in osteoblast cultures was also performed. Our data suggest that the origin of the gut microbiota inoculum plays a major role in the viability of osteoblasts. The treatments using P. ostreatus mushroom-powder and G. lucidum mushroom-extract had positive effects based on gut microbiota and SCFA analyses. Both mushrooms exhibited lower RANKL levels compared to controls, whereas their extracts tended to enhance the osteoblastic activity. In conclusion, mushrooms that are rich in beta-glucans may exert beneficial in vitro effects on bone physiology by alterations in the gut microbiota and/or SCFA production.

Priming of intestinal cytoprotective genes and antioxidant capacity by dietary phytogenic inclusion in broilers.

The potential of a phytogenic premix (PP) based on ginger, lemon balm, oregano and thyme to stimulate the expression of cytoprotective genes at the broiler gut level was evaluated in this study. In particular, the effects of PP inclusion levels on a selection of genes related to host protection against oxidation (catalase [CAT], superoxide dismutase 1 [SOD1], glutathione peroxidase 2 [GPX2], heme oxygenase 1 [HMOX1], NAD(P)H quinone dehydrogenase 1 [NQO1], nuclear factor (erythroid-derived 2)-like 2 [Nrf2] and kelch like ECH associated protein 1 [Keap1]), stress (heat shock 70 kDa protein 2 [HSP70] and heat shock protein 90 alpha family class A member 1 [HSP90]) and inflammation (nuclear factor kappa B subunit 1 [NF-κB1], Toll-like receptor 2 family member B (TLR2B) and Toll-like receptor 4 [TLR4]) were profiled along the broiler intestine. In addition, broiler intestinal segments were assayed for their total antioxidant capacity (TAC). Depending on PP inclusion level (i.e. 0, 750, 1,000 and 2,000 mg/kg diet) in the basal diets, 1-d-old Cobb broiler chickens (n = 500) were assigned into the following 4 treatments: CON, PP-750, PP-1000 and PP-2000. Each treatment had 5 replicates of 25 chickens with ad libitum access to feed and water. Data were analyzed by ANOVA and means compared using Tukey's honest significant difference (HSD) test. Polynomial contrasts tested the linear and quadratic effect of PP inclusion levels. Inclusion of PP increased (P ≤ 0.05) the expression of cytoprotective genes against oxidation, except CAT. In particular, the cytoprotective against oxidation genes were up-regulated primarily in the duodenum and the ceca and secondarily in the jejunum. Most of the genes were up-regulated in a quadratic manner with increasing PP inclusion level with the highest expression levels noted in treatments PP-750 and PP-1000 compared to CON. Similarly, intestinal TAC was higher in PP-1000 in the duodenum (P = 0.011) and the ceca (P = 0.050) compared to CON. Finally, increasing PP inclusion level resulted in linearly reduced (P ≤ 0.05) expression of NF-κB1, TLR4 and HSP70, the former in the duodenum and the latter 2 in the ceca. Overall, PP inclusion consistently up-regulated cytoprotective genes and down-regulated stress and inflammation related ones. The effect is dependent on PP inclusion level and the intestinal site. The potential of PP to beneficially prime bird cytoprotective responses merit further investigation under stress-challenge conditions.

Effects of Rich in Β-Glucans Edible Mushrooms on Aging Gut Microbiota Characteristics: An In Vitro Study.

Alterations of gut microbiota are evident during the aging process. Prebiotics may restore the gut microbial balance, with ß-glucans emerging as prebiotic candidates. This study aimed to investigate the impact of edible mushrooms rich in ß-glucans on the gut microbiota composition and metabolites by using in vitro static batch culture fermentations and fecal inocula from elderly donors (n = 8). Pleurotus ostreatus, P. eryngii, Hericium erinaceus and Cyclocybe cylindracea mushrooms derived from various substrates were examined. Gut microbiota composition (quantitative PCR (qPCR)) and short-chain fatty acids (SCFAs; gas chromatography (GC)) were determined during the 24-h fermentation. P. eryngii induced a strong lactogenic effect, while P. ostreatus and C. cylindracea induced a significant bifidogenic effect (p for all <0.05). Furthermore, P. eryngii produced on wheat straw and the prebiotic inulin had comparable Prebiotic Indexes, while P. eryngii produced on wheat straw/grape marc significantly increased the levels of tested butyrate producers. P. ostreatus, P. eryngii and C. cylindracea had similar trends in SCFA profile; H. erinaceus mushrooms were more diverse, especially in the production of propionate, butyrate and branched SCFAs. In conclusion, mushrooms rich in ß-glucans may exert beneficial in vitro effects in gut microbiota and/or SCFAs production in elderly subjects.

Dietary probiotic form modulates broiler gut microbiota indices and expression of gut barrier genes including essential components for gut homeostasis.

The probiotic form (PF) type and its dietary administration in combination or not with avilamycin (AV) were investigated for their effects on broiler gut microbiota and expression of genes relevant for gut barrier and gut homeostasis. Depending on PF type (i.e. no addition, viable, inactivated) and AV addition (no/yes), 450 one-day-old Cobb male broilers were allocated in 6 treatments (CON, CON + A, ViP, ViP + A, InP and InP + A) according to a 3 × 2 factorial arrangement with 5 replicates of 15 broilers each for 42 days. Significant interactions (PPF × AV  ≤ 0.05) between PF and AV administration were shown for the ileal mucosa-associated bacteria, the caecal digesta Lactobacillus spp., the molar ratio of the sum of valeric, hexanoic and heptanoic acids and the gene expressions of ileal and caecal IgA and ileal claudin 1. Avilamycin suppressed ileal digesta Lactobacillus spp. (PAV  < 0.001) and caecal digesta Clostridium perfringens subgroup (PAV  = 0.018) and modulated the intestinal fermentation intensity and pattern. The viable PF had the higher levels of ileal digesta Bacteroides spp. (PPF  = 0.021) and caecal digesta Lactobacillus spp. (PPF  = 0.038) compared with the other two PF. Probiotic form modulated the microbial metabolic activity in the ileum and caeca with the viable PF being the most noteworthy in terms of effects regarded as beneficial. Furthermore, the viable PF resulted in reduced expression of caecal Toll-like receptors TLR2B (PPF  = 0.026) and TLR4 (PPF  = 0.011) and transcription factor NFΚΒ1 (PPF  = 0.002), which could be considered as essential for limiting inflammation and preserving gut homeostasis. In conclusion, under non-challenge conditions, probiotic function was shown to depend on PF type and to a lesser degree on co-administration with AV. The importance of probiotic viability for the beneficial modulation of important gut components towards a reduced state of physiological inflammation has been highlighted.

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 (P CT = 0.007) and digesta Bifidobacterium (P CT < 0.001), as well as lower levels of total bacteria (P CT = 0.004) and Clostridia clusters I, IV and XIVa (P CT ≤ 0.05), compared with maize-fed ones. In addition, cereal type gave differences in fermentation intensity (P CT = 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) (P PFA = 0.023) and MUC2 (P PFA = 0.001) genes, and at 150 mg/kg decreased cecal TLR2 (P PFA = 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.

Modulation of broiler gut microbiota and gene expression of Toll-like receptors and tight junction proteins by diet type and inclusion of phytogenics.

This study evaluated the effect of reduced dietary energy (ME) and crude protein (CP) levels along with inclusion of a phytogenic feed additive (PFA) on gut microbiota composition and gene expression of Toll-like receptor(s) (TLR), tight junction proteins, and inflammatory cytokines expressed in secondary lymphoid organs. Depending on dietary ME and CP level down regulation and the inclusion or not of PFA at 125 mg/kg diet, 450 one-day-old male broilers were allocated in the following 6 treatments for 42 D according to a 3 × 2 factorial design: A: diet formulated optimally to meet broiler nutrient requirements; 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. Diet type and PFA supplementation were shown to affect mostly the mucosa-associated microbiota compared to the luminal ones. Ileal mucosa-associated total bacteria (PD= 0.005), Lactobacillus spp. (PD= 0.003), and Clostridium cluster XIVa (PD= 0.009) were affected by diet type with broilers fed diet B having lower levels compared to broilers fed diets A or C. Moreover, diet type affected cecal mucosa-associated Lactobacillus spp. (PD= 0.002) with broilers fed diet C having lower levels compared to broilers fed diets A or B. Supplementation with PFA resulted in higher levels of cecal mucosa-associated Bacteroides (PP= 0.031), Clostridium cluster IV (PP= 0.007), and Clostridium cluster XIVa (PP= 0.039). Diet type affected TLR2 (PD= 0.046) and claudin 5 (PD= 0.027) in cecal epithelium. Lower TLR2 (PP= 0.021) and higher zonula occludens 2 (PP= 0.031) relative gene expressions were seen in ileal epithelium following PFA supplementation. Moreover, in cecal epithelium, PFA supplementation resulted in lower TLR2 (PP < 0.001) and higher zonula occludens 2 (PP= 0.009), claudin 5 (PP= 0.005) and occludin (PP= 0.039) relative gene expressions. There were no significant diet type and PFA effects on cytokines in secondary lymphoid organs, except for a dietary effect on transforming growth factor beta 4 (PD= 0.023) in cecal tonsils. In conclusion, PFA inclusion beneficially modulated elements of gut microbiota, Toll-like signaling molecules and gut tight junction genes.

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 (P O × A = 0.028) compared with AV. In addition, ileal digesta levels of Clostridium perfringens subgroup were decreased by avilamycin (P A = 0.045). Inclusion of OABF stimulated the activity of microbial glycolytic enzymes, whereas avilamycin resulted in lower acetate (P A = 0.021) and higher butyrate (P A = 0.010) molar ratios. Expression of ZO1 and CLDN5 was down-regulated by both OABF (P O = 0.016 and P O = 0.003, respectively) and avilamycin (P A = 0.016 and P A = 0.001, respectively). In addition, CLDN1 was down-regulated in AV compared with CON (P O × A = 0.012). Furthermore, OABF down-regulated MUC2 (P O = 0.027), whereas avilamycin down-regulated nuclear factor kappa B subunit 1 (NFKB1) (P A = 0.024), toll-like receptor 2 family member B (TLR2B) (P A = 0.011) and toll-like receptor 4 (TLR4) (P A = 0.014) expression. In the ceca, OABF inclusion increased digesta levels of Clostridium coccoides (P O = 0.018) and Clostridium leptum (P O = 0.040) subgroups, while it up-regulated MUC2 expression (P O = 0.014). Avilamycin (P A = 0.044) and interaction (P O × 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.

Adherence to the Mediterranean diet is associated with the gut microbiota pattern and gastrointestinal characteristics in an adult population.

This study aimed to explore the potential associations of adherence to the Mediterranean diet with gut microbiota characteristics and gastrointestinal symptomatology in an adult population. Other long-term dietary habits (e.g. consumption of snacks and junk food or stimulant intake) were also evaluated in terms of the gut microbiota profile. Participants (n 120) underwent anthropometric, dietary, physical activity and lifestyle evaluation. Adherence to the Mediterranean diet was assessed using a Mediterranean diet score, the MedDietScore, and subjects were classified into three tertiles according to individual adherence scoring. Gut microbiota composition was determined using quantitative PCR and plate-count techniques, and faecal SCFA were analysed using GC. Gastrointestinal symptoms were also evaluated. Participants with a high adherence to the Mediterranean diet had lower Escherichia coli counts (P=0·022), a higher bifidobacteria:E. coli ratio (P=0·025), increased levels and prevalence of Candida albicans (P=0·039 and P=0·050, respectively), greater molar ratio of acetate (P=0·009), higher defaecation frequency (P=0·028) and a more pronounced gastrointestinal symptomatology compared with those reporting low adherence. A lower molar ratio of valerate was also observed in the case of high adherence to the Mediterranean diet compared with the other two tertiles (P for trend=0·005). Positive correlations of MedDietScore with gastrointestinal symptoms, faecal moisture, total bacteria, bifidobacteria:E. coli ratio, relative share of Bacteroides, C. albicans and total SCFA, as well as negative associations with cultivable E. coli levels and valerate were indicated. Fast food consumption was characterised by suppressed representation of lactobacilli and butyrate-producing bacteria. In conclusion, our findings support a link between adherence to the Mediterranean diet and gut microbiota characteristics.

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

BACKGROUND: Live commensal intestinal bacteria are present in the peripheral blood where they can induce inflammation. OBJECTIVE: To evaluate the intestinal bacteria composition and translocation of bacteria in IBD. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

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.