In ovo Injection of a Galacto-Oligosaccharide Prebiotic in Broiler Chickens Submitted to Heat-Stress: Impact on Transcriptomic Profile and Plasma Immune Parameters.
ABSTRACT: This study investigated the effects of a galactooligosaccharide (GOS) prebiotic in ovo injected on intestinal transcriptome and plasma immune parameters of broiler chickens kept under thermoneutral (TN) or heat stress (HS) conditions. Fertilized Ross 308 eggs were injected in ovo with 0.2 mL physiological saline without (control, CON) or with 3.5 mg of GOS (GOS). Three-hundred male chicks/injection treatment (25 birds/pen) were kept in TN or HS (30 °C) conditions during the last growing phase, in a 2 × 2 factorial design. At slaughter, from 20 birds/injection group (half from TN and half from HS), jejunum and cecum were collected for transcriptome analysis, and plasma was collected. No differences in plasma parameters (IgA and IgG, serum amyloid) and no interaction between injection treatment and environment condition were found. GOS-enriched gene sets related to energetic metabolism in jejunum, and to lipid metabolism in cecum, were involved in gut barrier maintenance. A homogeneous reaction to heat stress was determined along the gut, which showed downregulation of the genes related to energy and immunity, irrespective of in ovo treatment. GOS efficacy in counteracting heat stress was scarce after ten days of environmental treatment, but the in ovo supplementation modulates group of genes in jejunum and cecum of broiler chickens.
Project description:Galactooligosaccharides (GOS) that are delivered in ovo improve intestinal microbiota composition and mitigate the negative effects of heat stress in broiler chickens. Hubbard hybrids are slow-growing chickens with a high resistance to heat. In this paper, we determined the impact of GOS delivered in ovo on slow-growing chickens that are challenged with heat. The experiment was a 2 × 2 × 2 factorial design. On day 12 of incubation, GOS (3.5 mg/egg) was delivered into the egg (n = 300). Controls (C) were mock-injected with physiological saline (n = 300). After hatching, the GOS and C groups were split into thermal groups: thermoneutral (TN) and heat stress (HS). HS (30 °C) lasted for 14 days (days 36-50 post-hatching). The spleen (n = 8) was sampled after acute (8.5 h) and chronic (14 days) HS. The gene expression of immune-related (IL-2, IL-4, IL-6, IL-10, IL-12p40, and IL-17) and stress-related genes (HSP25, HSP90AA1, BAG3, CAT, and SOD) was detected with RT-qPCR. Chronic HS up-regulated the expression of the genes: IL-10, IL-12p40, SOD (p < 0.05), and CAT (p < 0.01). GOS delivered in ovo down-regulated IL-4 (acute p < 0.001; chronic p < 0.01), IL-12p40, CAT and SOD (chronic p < 0.05). The obtained results suggest that slow-growing hybrids are resistant to acute heat and tolerant to chronic heat, which can be supported with in ovo GOS administration.
Project description:Intestinal mucosa is the interface between the microbial content of the gut and the host's milieu. The goal of this study was to modulate chicken intestinal microflora by in ovo stimulation with galactooligosaccharides (GOS) prebiotic and to demonstrate the molecular responses of the host. The animal trial was performed on meat-type chickens (Ross 308). GOS was delivered by in ovo injection performed into the air cell on day 12 of egg incubation. Analysis of microbial communities and mucosal gene expression was performed at slaughter (day 42 post-hatching). Chyme (for DNA isolation) and intestinal mucosa (for RNA isolation) from four distinct intestinal segments (duodenum, jejunum, ileum, and caecum) was sampled. The relative abundance of Bifidobacterium spp. and Lactobacillus spp. in DNA isolated from chyme samples was determined using qPCR. On the host side, the mRNA expression of 13 genes grouped into two panels was analysed with RT-qPCR. Panel (1) included genes related to intestinal innate immune responses (IL-1?, IL-10 and IL-12p40, AvBD1 and CATHL2). Panel (2) contained genes involved in intestinal barrier function (MUC6, CLDN1 and TJAP1) and nutrients sensing (FFAR2 and FFAR4, GLUT1, GLUT2 and GLUT5). GOS increased the relative abundance of Bifidobacterium in caecum (from 1.3% to 3.9%). Distinct effects of GOS on gene expression were manifested in jejunum and caecum. Cytokine genes (IL-1?, IL-10 and IL-12p40) were up-regulated in the jejunum and caecum of the GOS-treated group. Host defence peptides (AvBD1 and CATHL2) were up-regulated in the caecum of the GOS-treated group. Free fatty acid receptors (FFAR2 and FFAR4) were up-regulated in all three compartments of the intestine (except the duodenum). Glucose transporters were down-regulated in duodenum (GLUT2 and GLUT5) but up-regulated in the hindgut (GLUT1 and GLUT2). In conclusion, GOS delivered in ovo had a bifidogenic effect in adult chickens. It also modulated gene expression related to intestinal immune responses, gut barrier function, and nutrient sensing.
Project description:There is increasing interest in the use of natural antioxidant supplements in poultry diets as protection against the adverse effects of heat stress. The potential protective effect of boldo (Peumus boldus molina) leaf extract, which have antioxidant activity, were investigated against the harmful effects of heat stress in two broiler strains. Arbor Acres (AA) and Avian-48 (AV) chicks were divided into thermoneutral (TN) and heat stress (HS) groups and treated with 1 g boldo leaf extract/4 L drinking water during the heat stress period. HS reduced growth performance in both strains. The phagocytic index, phagocytic activity, and eosinophil and lymphocytes counts were significantly elevated in TN and HS AV birds but not altered in AA birds. Boldo extract treatment partially eliminated the previous negative impacts of heat stress. AA chicks were better able to withstand HS than AV chicks. Serum concentrations of total lipids and cholesterol were reduced in HS birds of both strains. Malondialdehyde, superoxide dismutase, and glutathione peroxidase levels were elevated but restored with the administration of boldo leaf extract in HS birds of both strains. Economic parameters were negatively affected by HS but restored to values close to those of the control group in boldo-treated HS birds. In conclusion, the administration of boldo leaf extract in drinking water was effective in neutralizing the harmful effects of heat stress on growth performance, blood indices, and economic parameters and improved the antioxidant defense system in heat-stressed birds.
Project description:Accumulating evidence has revealed the dysbiosis of gut/fecal microbiota induced by heat stress (HS) in mammals and poultry. However, the effects of HS on microbiota communities in different intestinal segments of Cherry-Valley ducks (a widely used meat-type breed) and their potential associations with growth performances, fat deposition, intestinal morphology, and antioxidant capacity have not been well evaluated yet. In this study, room temperature (RT) of 25°C was considered as control, and RT at 32°C for 8 h per day was set as the HS treatment. After 3 weeks, the intestinal contents of jejunum, ileum, and cecum were harvested to investigate the microbiota composition variations by 16S ribosomal RNA amplicon sequencing. And the weight gain, adipose indices, intestinal morphology, and a certain number of serum biochemical parameters were also measured and analyzed. The results showed the microbial species at different levels differentially enriched in duck jejunum and cecum under HS, while no significant data were observed in ileum. HS also caused the intestinal morphological changes (villus height and the ratio of villus height to crypt depth) and the reductions of growth speed (daily gain), levels of serum triglyceride (TG) and total cholesterol, and antioxidant activity (higher malondialdehyde (MDA) content and lower total antioxidant). The higher abdominal fat content and serum glucose level were also observed in HS ducks. The Spearman correlation analysis indicated that in jejunum the phyla Firmicutes and Proteobacteria were associated with average daily gain, feed/gain, serum TG and MDA levels, and villus height/crypt depth (P < 0.05). The phylum Firmicutes and genus Acinetobacter were significantly associated with fat deposition and serum glucose level (P < 0.05). The genus Lactobacillus was positively associated with serum total antioxidant (P < 0.05), while some other microbial species were found negatively associated, including order Pseudomonadales, genera Acinetobacter, and unidentified_Mitochondria. However, no significant correlations were observed in cecum. These findings imply the potential roles of duck gut microbiota in the intestinal injuries, fat deposition, and reductions of growth speed and antioxidant capacity caused by HS, although the molecular mechanisms requires further investigation.
Project description:Synbiotics are synergistic combinations of prebiotics and probiotics. In chickens, synbiotics can be delivered in ovo to expedite colonization of the gut by beneficial bacteria. We therefore aimed to design synbiotics in vitro and validate them in broiler chickens upon in ovo delivery. The probiotic components of the synbiotics were Lactobacillus salivarius and Lactobacillus plantarum. Their growth was assessed in MRS medium supplemented with different prebiotics. Based on in vitro results (hatchability and growth curve), two synbiotics were designed: S1 -Lactobacillus salivarius with galactooligosaccarides (GOS) and S2 -Lactobacillus plantarum with raffinose family oligosaccharides (RFO). These synbiotics were delivered to Cobb broiler chicken embryos on day 12 of incubation at optimized doses (105 cfu egg-1 of probiotic, 2 mg egg-1 of prebiotic). Post hatching, 2,400 roosters were reared (600 individuals group-1 divided into eight replicate pens). Microbial communities were analyzed in ileal and cecal digesta on day 21 using FISH. Gene expression analysis (IL1?, IL4, IL6, IL8, IL12, IL18, IFN?, and IFN?) was performed on days 7, 14, 21, and 42 for the spleen and cecal tonsils with RT-qPCR. Body weight and feed intake of the roosters did not differ by the treatments. Microbial populations of Lactobacillus spp. and Enterococcus spp. in the ileum were higher in S1 and S2 than in the control. In the cecum, the control had the highest bacterial counts. S1 caused significant up-regulation of IL6, IL18, IL1?, IFN?, and IFN? in the spleen on day 21 and IL1? on day 7 (P < 0.05). In cecal tonsils, S1 caused significant down-regulation of IL12, IL8, and IL1? on day 42 and IFN? on day 14 (P < 0.05). S2 did not elicit such patterns in any tissues investigated. Thus, we demonstrate that divergent effects of synbiotics in broiler chickens were reflected in in vitro tests.
Project description:Heat stress (HS) adversely affects growth performance and inflicts heavy economic losses to the poultry industry. There is, therefore, a critical need to identify new alternative strategies to alleviate the negative effects induced by HS. The tropic medicinal plant, Morinda citrifolia (Noni), is being used in livestock nutrition, however the literature is limited and conflicting for its impact on growth performance. The present study aimed to determine the effect of Noni on feeding and drinking behavior as well as on the hypothalamic expression of stress- and metabolic-related genes in broiler chickens exposed to acute HS. A total of 480 1 day-old male broiler chicks were randomly assigned to 12 controlled environmental chambers. Birds were subjected to two environmental conditions (TN, 25°C vs. HS, 35°C for 2 h) and fed two diets (control vs. 0.2% Noni) in a 2 × 2 factorial design. Feed intake and core body temperature (BT) were recorded during HS period. Blood was collected and hypothalamic tissues were harvested for target gene and protein analyses. Acute HS-broilers exhibited higher BT (~1°C), spent less time eating with a significant decrease in feed intake, and spent more time drinking along with higher drinking frequency compared to those maintained under TN conditions. Although Noni supplementation did not improve feed intake, it significantly delayed (~30 min) and reduced the BT-induced by HS. At molecular levels and under HS conditions, Noni supplementation down regulated the hypothalamic expression of HSP90 and its related transcription factors HSF1, 2, and 4, increased orexin mRNA levels, and decreased the phosphorylation levels of AMPK?1/2Thr172 and mTORSer2481. Together, these data indicated that Noni supplementation might modulate HS response in broilers through central orexin-AMPK-mTOR pathways.
Project description:High ambient temperatures negatively affect the human well-being as well as animal welfare and production. The gastrointestinal tract is predominantly responsive to heat stress. The currently available information about the multifaceted response to heat stress within different parts of the intestine is limited, especially in avian species. Hence, this study aims to evaluate the heat stress-induced sequence of events in the intestines of chickens. Furthermore, the gut health-promoting effect of dietary galacto-oligosaccharides (GOS) was investigated in these heat stress-exposed chickens. Chickens were fed a control diet or diet supplemented with 1% or 2.5% GOS (6 days) prior to and during a temperature challenge for 5 days (38-39°C, 8h per day). The parameters measured in different parts of the intestines included the genes (qPCR) HSF1, HSF3, HSP70, HSP90, E-cadherin, claudin-1, claudin-5, ZO-1, occludin, TLR-2, TLR-4, IL-6, IL-8, HO-1, HIF-1?) and their associated proteins HSP70, HSP90 and pan-cadherin (western blots). In addition, IL-6 and IL-8 plasma concentrations were measured by ELISA. In the jejunum, HSF3, HSP70, HSP90, E-cadherin, claudin-5, ZO-1, TLR-4, IL-6 and IL-8 mRNA expression and HSP70 protein expression were increased after heat stress exposure and a more pronounced increase in gene expression was observed in ileum after heat stress exposure, and in addition HSF1, claudin-1 and HIF-1? mRNA levels were upregulated. Furthermore, the IL-8 plasma levels were decreased in chickens exposed to heat stress. Interestingly, the heat stress-related effects in the jejunum were prevented in chickens fed a GOS diet, while dietary GOS did not alter these effects in ileum. In conclusion, our results demonstrate the differences in susceptibility to heat stress along the intestine, where the most obvious modification in gene expression is observed in ileum, while dietary GOS only prevent the heat stress-related changes in jejunum.
Project description:Palm kernel cake (PKC) is the main byproduct from the palm oil industry in several tropical countries that contains considerable amounts of oligosaccharide. We earlier demonstrated beneficial prebiotic effects of oligosaccharides extract of PKC (OligoPKC) in starter and finisher broiler birds. This study was envisaged to elucidate the effects of in ovo and/or oral administration of the OligoPKC on prenatal and post-hatched broiler chicks. A total of 140 broiler (Cobb500) eggs were randomly divided into two groups (n = 70 each), and on day 12 of incubation, eggs in one group received in ovo injection of 0.1 mL (containing 20 mg) of OligoPKC, while those in the other group received 0.1 mL of saline (placebo) solution. Of these in ovo placebo or OligoPKC injected eggs, after hatching, six chicks from each group were sampled for day-one analysis, while 48 chicks from each group were randomly allocated to two dietary regimes involving either no feeding or feeding of OligoPKC through basal diet for a 14 days experiment forming the experimental groups as: (i) saline-injected (Control, C), (ii) OligoPKC-injected (PREBovo), (iii) saline-injected, but fed 1% OligoPKC (PREBd), and (iv) OligoPKC-injected and also 1% OligoPKC (PREBovo+d). In ovo injection of prebiotic OligoPKC had no effect on body weight and serum immunoglobulins concentrations of day old chicks, except for IgG, which was increased significantly (P<0.05). Body weight and feed conversion ratio of 14 days old chicks were neither affected by in ovo injection nor feeding of OligoPKC. However, populations of cecal total bacteria and major beneficial bacteria of the chicks were markedly enhanced by feeding of OligoPKC (PREBd and PREBovo+d > C and PREBovo), but lesser influenced by in ovo OligoPKC injection. Irrespective of its prior in ovo exposure, chicks fed OligoPKC diets had lower population of pathogenic bacteria. Overall serum immunoglobulin status of birds was improved by feeding of OligoPKC but in ovo OligoPKC injection had minor effect on that. In most cases, in ovo OligoPKC injection and feeding of OligoPKC reduced the expression of nutrient transporters in the intestine and improved antioxidant capacity of liver and serum. It is concluded that in ovo injection of OligoPKC increased IgG production and antioxidant capacity in serum and liver of prenatal chicks and had limited carrying-over effects on the post-hatched chicks comparing to the supplementary feeding of OligoPKC.
Project description:Study objectives were to determine the effects of chromium (Cr) propionate (Cr propionate 0.04%; 0.5 g/kg of feed to deliver 200 parts per billion Cr/d; KemTRACE Cr, Kemin Industries, Inc., Des Moines, IA) on growth performance, metabolism, and health biomarkers in heat-stressed and nutrient-restricted pigs. Crossbred barrows (n = 96; 105 ± 1 kg BW) were enlisted in an experiment conducted in two replicates, blocked by initial BW, and randomly assigned to one of six dietary-environmental treatments: (i) thermoneutral (TN) and fed ad libitum a control diet (TNCtl), (ii) TN and fed ad libitum a Cr supplemented diet (TNCr), (iii) TN and pair-fed a control diet (PFCtl), (iv) TN and pair-fed a Cr supplemented diet (PFCr), (v) heat stress (HS) and ad libitum fed a control diet (HSCtl), or (vi) HS and ad libitum fed a Cr supplemented diet (HSCr). The study consisted of three experimental periods (P). During P0 (5 d), all pigs were housed in TN conditions (21.3 ± 0.1 °C, 56.8 ± 0.3% relative humidity [RH]) and fed the control diet ad libitum. During P1 (5 d), pigs were fed their respective dietary treatments ad libitum and kept in TN conditions. During P2 (35 d), HSCtl and HSCr-treated pigs were fed ad libitum and exposed to progressive cyclical HS conditions (27 to 31 °C, 50 ± 0.3% RH), while TNCtl, TNCr, PFCtl, and PFCr pigs remained in TN conditions and were fed ad libitum or pair-fed to their respective HSCtl and HSCr counterparts to eliminate the confounding effects of dissimilar feed intake. Overall, HS pigs had increased (P < 0.01) rectal temperature, skin temperature, and respiration rate (0.3 °C, 3.8 °C, and 32 breaths per minute, respectively) relative to TN pigs. Overall, HS decreased ADFI and ADG (20 and 21%, respectively; P < 0.01) compared with TN controls. Final BW tended to be increased in HSCr (2.7 kg, P = 0.06) compared with HSCtl pigs. Similarly, ADG tended to be increased during P2 in HSCr relative to HSCtl-treatment (0.77 vs. 0.72 kg/d; P = 0.10). There were no effects of Cr on most production parameters, but ADFI tended to be increased in Cr relative to Ctl-fed pigs (3.19 vs. 3.09 kg/d; P = 0.08). No effects of Cr supplementation were detected on circulating glucose, insulin, NEFA, cholesterol, triglycerides, or lipopolysaccharide binding protein. However, blood neutrophils were increased in HSCr (37%; P < 0.01) relative to HSCtl pigs. In summary, these results suggest Cr supplementation may benefit growth performance during HS.
Project description:Rats (n=24) were fed ad libitum diets containing either ground endophyte-free (E-) seed or endophyte–infected (E+) seed for five days at thermoneutrality (21°C) or heat stress (32 C) for 21 days. Rats with intraperitonial transmitters were used with core temperature (Tc) and general activity measured every 5 minutes during the study. At treatment end, the liver was removed, weighed and frozen in liquid nitrogen. RNA was extracted from liver samples, converted to cDNA, and hybridized with the printed oligonucleotide slides. Treatments consisted of four treatment groups, E-TN, E+TN, E-HS, E+HS. E-TN IS control at thermoneutral for 26 days , E+TN toxin fed group at thermoneutral for 26 days; E-HS control at thermoneutral for 5 days and heat treatment for 21 days, E+HS is toxin fed group at thermoneutral for 5 days and heat stress for 21 days. Keywords: Stress response Overall design: Rats (n=24) were divided into two groups and fed with either E- or E+ diet for 5 days under TN conditions. At the end of TN period, each group was further subdivided into two groups each. Treatments consisted of four treatment groups, E-TN, E+TN, E-HS, E+HS. E-TN IS control at thermoneutral for 26 days , E+TN toxin fed group at thermoneutral for 26 days; E-HS control at thermoneutral for 5 days and heat treatment for 21 days, E+HS is toxin fed group at thermoneutral for 5 days and heat stress for 21 days.