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Effects of thymol and carvacrol supplementation on intestinal integrity and immune responses of broiler chickens challenged with Clostridium perfringens.

ABSTRACT: BACKGROUND:Necrotic enteritis caused by Clostridium perfringens infection leads to serious economic losses in the global poultry production. In the present study, we investigated the protective effects of essential oils (EO, which contained 25 % thymol and 25 % carvacrol as active components) supplementation on growth performance, gut lesions, intestinal morphology, and immune responses of the broiler chickens infected with C. perfringens. A total of 448 1-day-old male broiler chicks were allocated into eight treatment groups following a 4?×?2 factorial arrangement with four dietary EO dosages (0, 60, 120, or 240 mg/kg) and two infection status (with or without C. perfringens challenge from d 14 to 20). RESULTS:The challenge did not impair the growth performance of birds, but induced gut lesions and increased crypt depth in the ileum (P???0.05). It also down-regulated the claudin-1 and occludin mRNA expression (P???0.05), up-regulated the mRNA expression of interleukin-1? (P???0.05), tended to increase the toll-like receptor (TLR) 2 mRNA expression (P?

SUBMITTER: Du E

PROVIDER: S-EPMC4802587 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Effects of thymol and carvacrol supplementation on intestinal integrity and immune responses of broiler chickens challenged with Clostridium perfringens.

Du Encun E Wang Weiwei W Gan Liping L Li Zhui Z Guo Shuangshuang S Guo Yuming Y

Journal of animal science and biotechnology 20160322

<h4>Background</h4>Necrotic enteritis caused by Clostridium perfringens infection leads to serious economic losses in the global poultry production. In the present study, we investigated the protective effects of essential oils (EO, which contained 25 % thymol and 25 % carvacrol as active components) supplementation on growth performance, gut lesions, intestinal morphology, and immune responses of the broiler chickens infected with C. perfringens. A total of 448 1-day-old male broiler chicks wer ...[more]

PMID: 27006768

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Project description:Our previous reports suggested that Dietary l-arginine supplementation attenuated gut injury of broiler chickens infected with Clostridium perfringens by enhancing intestinal immune responses, absorption and barrier function, but its effect on the gut microbiome of broiler chickens remains unclear. This experiment aimed at evaluating the effects of Dietary l-arginine supplementation on the gut bacterial community composition and function of broiler chickens challenged with C. perfringens. In total, 105 1-day-old male Arbor Acres broiler chickens were assigned to three groups: Control (CTL), C. perfringens-challenged (CP), and C. perfringens-challenged and fed diet supplemented with 0.3% l-arginine (ARGCP) groups. The challenge led to macroscopic and histomorphological gut lesions, decreased villus height and increased the number of Observed species, Shannon, Chao1 and ACE indices of ileal microbiota, whereas l-arginine addition reversed these changes. Moreover, the three treatments harbored distinct microbial communities (ANOSIM, P < 0.05). At the genus level, 24 taxa (e.g., Nitrosomonas spp., Coxiella spp., Ruegeria spp., and Thauera spp.) were significantly more abundant in CP group than in CTL group (P < 0.05), whereas the levels of 23 genera of them were significantly decreased by l-arginine supplementation (P < 0.05). The abundances of only 3 genera were different between CTL and ARGCP groups (P < 0.05). At the species level, the challenge promoted the relative abundance of Nitrospira sp. enrichment culture clone M1-9, Bradyrhizobium elkanii, Nitrospira bacterium SG8-3, and Pseudomonas veronii, which was reversed by l-arginine supplementation (P < 0.05). Furthermore, the challenge decreased the levels of Lactobacillus gasseri (P < 0.05). Predictive functional profiling of microbial communities by PICRUSt showed that compared with CP group, ARGCP group had enriched pathways relating to membrane transport, replication and repair, translation and nucleotide metabolism and suppressed functions corresponding to amino acid and lipid metabolisms (P < 0.05). The relative abundances of KEGG pathways in l-arginine-fed broilers were almost equal to those of the controls. In conclusion, l-arginine alleviated the gut injury and normalized the ileal microbiota of C. perfringens-challenged chickens to resemble that of unchallenged controls in terms of microbial composition and functionality.

Project description:The primary cause of necrotic enteritis (NE) disease in chickens is the NetB-positive Clostridium perfringens bacterium. Many factors are known to affect the severity of NE in the challenge models of broiler chickens, and one of these factors is the virulence of C. perfringens strain. This study was conducted to evaluate the effect of 2 pathogenic C. perfringens strains in a NE challenge model on gut health and mRNA expression of genes encoding apoptosis, tight junction, immunity, and nutrient transporters in broilers. Day-old Ross-308 male broilers (n = 468) were allocated in a 2 × 3 factorial arrangement of treatments with in-feed antibiotics (no or yes) and challenge (Non, C. perfringens strain NE18, and C. perfringens strain NE36) as the factors. The birds in the challenged groups were inoculated with Eimeria species on day 9 and with a fresh suspension of C. perfringens NE18 or NE36 on day 14 and 15. Sample collection was performed on 2 birds of each pen on day 16. Necrotic enteritis challenge, impaired feed conversion ratio during day 0 to 16 compared with the control group where the effect of the NE36 challenge was more severe than that with NE18 (P < 0.001). The mRNA expression of mucin-2, immunoglobulin-G, occludin (P < 0.001), and tight junction protein-1 (P < 0.05) genes were downregulated in both challenged groups compared with the nonchallenged counterparts. Antibiotic supplementation, on the other hand, increased weight gain, and feed intake in all challenged birds (P < 0.01), but upregulated mucin-5ac and alanine, serine, cysteine, and threonine transporter-1 (P < 0.05) only in the NE18 challenged birds. The challenge with NE36 significantly upregulated caspase-8 and claudin-1 (P < 0.001), but downregulated glucose transporter-2 (P < 0.001) compared with the NE18 challenge. These results suggest that NE challenge is detrimental to the performance of broilers through compromised intestinal health, and different C. perfringens strains can affect the severity of the disease through modulating the expression of intestinal genes encoding proteins responsible for apoptosis, gut integrity, immunity, mucus production, and nutrient transporters.

Project description:Preventative effects of Lactobacillus fermentum and Bacillus coagulans against Clostridium perfringens infection in broilers have been well-demonstrated. The present study was conducted to investigate the modulation of these two probiotics on intestinal immunity and microbiota of C. perfringens-challenged birds. The 336 one-day-old broilers were assigned to four groups with six replicates in each group. Birds in the control were unchallenged and fed a basal diet, and birds in the three challenged groups were dietary supplemented with nothing (Cp group), 1 × 109 CFU/kg of L. fermentum (Lf_Cp group), or 1 × 1010 CFU/kg of B. coagulans (Bc_Cp group). Challenge was performed from days 14 to 20, and samples were collected on days 21 and 28. Challenge upregulated interleukin (IL)-1β and transforming growth factor (TGF)-β4 mRNA expression in jejunum on day 21, which was downregulated by B. coagulans and L. fermentum, respectively (P < 0.05). Both probiotic groups upregulated jejunal IL-1β, interferon (IFN)-γ, IL-17, and TGF-β4 on day 28 as well as IFN-γ on day 21 (P < 0.05). The Bc_Cp group increased CD3+ T cell counts in the jejunal crypt on day 21 (P < 0.05). Challenge decreased the ileal ACE index on day 21 and cecal microbial richness on day 28, which were increased by probiotic treatments, and ileal bacterial richness decreased in the Bc_Cp group on day 28 (P < 0.05). Only ileal microbiota on day 21 was distinctly affected with an R-value at 0.3116 by ANOSIM analysis (P < 0.05). Compared with the control, ileal Firmicutes increased on day 21, and ileal Bacteroidetes and cecal Proteobacteria decreased on day 28 in challenged groups (P < 0.05). Challenge increased Romboutsia spp. in the ileum as well as unclassified f_Lachnospiraceae and Ruminococcus_torques group in the cecum, and decreased Lactobacillus spp. in the ileum on day 21, which were all conversely modulated by L. fermentum (P < 0.05). Challenge increased amino acid metabolism of ileal microbiota and membrane transport of cecal microbiota, and decreased amino acid metabolism of cecal microbiota on day 21, which were conversely regulated by both probiotics (P < 0.05). In conclusion, L. fermentum and B. coagulans attenuated the intestinal inflammation and microbial dysbiosis soon after C. perfringens challenge.

Project description:Necrotic enteritis toxin B (NetB)-producing Clostridium perfringens (CP) type A is the etiological agent of necrotic enteritis (NE) - an economically significant disease in broiler chickens. Understanding the immune response to CP infection in broiler chickens is becoming important to develop effective vaccines against NE. An experiment was conducted to determine the expression levels of selected cytokine genes in the intestine and cecal tonsil of CP-challenged broiler chickens. In a floor-pen housing, broiler chickens were randomly assigned to the following treatment groups: 1) bacitracin methylene disalicylate (BMD)-free control diet with no CP challenge (CX), 2) BMD-supplemented diet with no CP challenge (CM), 3) BMD-free control diet with CP challenge (PCX), or 4) BMD-supplemented diet with CP challenge (PCM). The establishment of CP infection was confirmed, with the treatment groups exposed to CP having a 1.5 to 2-fold higher CP levels (P < 0.05) compared to the non-exposed groups. On day 1 and 7 post-challenge, jejunal segments and cecal tonsils were collected from experimental chickens for quantitative real-time RT-PCR analysis to determine the expression levels of interleukin (IL)-1β, interferon-γ (IFN-γ), IL-2, IL-13, IL-17, IL-10, and transforming growth factor (TGF)-β genes. Levels of antibodies to CP recombinant proteins were also determined in the plasma of experimental chickens. Results indicated that on day 7 post-challenge, IL-1β (proinflammatory cytokine), IL-13 (Th2 cytokine), and IL-17 (Th17 cytokine) were upregulated (P < 0.05) in CP-challenged PCX and PCM treatments, compared to the unchallenged (control) CX and CM treatments. A reverse trend was observed for TGF-β (anti-inflammatory cytokine), while no change was observed in IFN-γ (Th1 cytokine). Levels of plasma antibodies (IgY) to CP recombinant proteins were higher in CP-challenged treatments (PCX and PCM; P < 0.05), compared to their corresponding controls (CX and CM). It was concluded that CP infection induced inflammatory response in the intestine of broiler chickens, and the mechanisms of inflammation are probably mediated via Th2 and Th17 cells.

Project description:Clostridium perfringens-induced necrotic enteritis (NE) is an economically important disease of broiler chickens. The present study evaluated the effect of C. perfringens on the intestinal histomorphometry, enteric microbial colonization, and host immune responses using 3 experimental NE reproduction methods. The experimental groups consisted of 1) unchallenged Control diet (corn-soybean meal), 2) Control diet + Eimera inoculation at d 11 followed by C. perfringens challenge at d 15 (ECp), 3) Wheat-based diet + C. perfringens challenge (WCp), and 4) Wheat-based diet + Eimeria inoculation followed by C. perfringens challenge (WECp). The results showed that chickens receiving ECp and WECp had reduced (P < 0.05) bird performance coupled with enteric gross lesions and epithelial damage at d 17 and 24 of age compared to unchallenged control birds. These ECp and WECp administered birds also had increased (P < 0.05) ileal colonization by clostridia and E. coli at d 17 and 24, while the resident Lactobacillus counts were reduced (P < 0.05) at d 24 of age. Furthermore, at d 24, jejunal transcription of IL-6, IL-10, annexin-A1 and IL-2 genes was upregulated (P < 0.05) in the ECp group, whereas the transcription of TNF receptor associated factor (TRAF)-3 gene was increased (P < 0.05) in WECp treated birds when compared to unchallenged control group. Additionally, stimulation of chicken splenocytes and cecal tonsilocytes with virulent C. perfringens bacilli or their secretory proteins resulted in a higher (P < 0.05) frequency of T cells and their upregulation of MHC-II molecule, as determined by flow cytometry. These findings suggest that C. perfringens, while inducing epithelial damage and changes in microbiota, can also trigger host immune responses. Furthermore, NE reproduction methods using coccidia with or without the wheat-based dietary predisposition seem to facilitate an optimal NE reproduction in broiler chickens and thus, may provide better avenues for future C. perfringens research.

Project description:Fusarium toxins are one of the most common contaminants in poultry diets. The co-occurrence of fumonisins (FUM) and deoxynivalenol (DON), even at a subclinical dose, negatively affects the growth performance, intestinal integrity and induce subclinical necrotic enteritis in broiler chickens. Loss of gut integrity can be expected to alter the intestinal microbiota’s composition. The objective of this study was to identify the effects of combined FUM and DON on the cecal microbiome profile and predicted metabolic functions and a short chain fatty acid profile in broilers challenged with Clostridium perfringens. A total of 240 1 day-old chicks were randomly assigned to two treatments: a control diet and the control diet with 3 mg/kg FUM + 4 mg/kg DON each with eight replications. All the birds were received cocci vaccine at d0. All birds in both treatment groups were challenged with C. perfringens 1 × 108 CFU via feed on d 19 and 20 to achieve 5% mortality. On d 35, the FUM and DON contaminated diet numerically (P = 0.06) decreased the body weight gain (BWG) by 84 g compared to the control group. The bacterial compositions of the cecal contents were analyzed by sequencing the V3–V4 region of the 16S rRNA gene. Overall, microbial richness and diversity increased (P < 0.02) during the studied period (d 21–35). Cecal contents of birds in the FUM + DON group had greater (P < 0.05) microbial evenness and diversity (Shannon index) compared to the control group. FUM + DON exposure decreased (P = 0.001) the relative abundance of Proteobacteria in the cecal content, compared to the control group. The combined FUM + DON significantly increased the relative abundance of the Defluviitaleaceae and Lachnospiraceae families (P < 0.05) but decreased the abundances of the Moraxellaceae and Streptococcaceae (P < 0.05) compared to the control group birds. At the genus level, FUM + DON exposure decreased (P < 0.05) Acinetobacter and Pseudomonas abundance and had a tendency (P = 0.08) to decrease Thermincola abundance compared to the control group. In the ileum, no NE-specific microscopic abnormalities were found; however, the tip of the ileal villi were compromised. The present findings showed that dietary FUM and DON contamination, even at subclinical levels, altered cecal microbial composition, dysregulated intestinal functions, and impaired the gut immune response, potentially predisposing the birds to necrotic enteritis.

Project description:The present study explored the potential effect of pterostilbene as a prophylactic treatment on the lipopolysaccharide (LPS)-induced intestinal injury of broiler chickens by monitoring changes in mucosal injury indicators, redox status, and inflammatory responses. In total, 192 one-day-old male Ross 308 broiler chicks were randomly divided into four groups. This trial consisted of a 2 × 2 factorial design with a diet factor (supplemented with 0 or 400 mg/kg pterostilbene from 1 to 22 d of age) and a stress factor (intraperitoneally injected with saline or LPS at 5.0 mg/kg BW at 21 da of age). The results showed that LPS challenge induced a decrease in BW gain (P < 0.001) of broilers during a 24-h period postinjection; however, this decrease was prevented by pterostilbene supplementation (P = 0.031). Administration of LPS impaired the intestinal integrity of broilers, as indicated by increased plasma diamine oxidase (DAO) activity (P = 0.014) and d-lactate content (P < 0.001), reduced jejunal villus height (VH; P < 0.001) and the ratio of VH to crypt depth (VH:CD; P < 0.001), as well as a decreased mRNA level of jejunal tight junction protein 1 (ZO-1; P = 0.002). In contrast, pterostilbene treatment increased VH:CD (P = 0.018) and upregulated the mRNA levels of ZO-1 (P = 0.031) and occludin (P = 0.024) in the jejunum. Consistently, pterostilbene counteracted the LPS-induced increased DAO activity (P = 0.011) in the plasma. In addition, the LPS-challenged broilers exhibited increases in nuclear accumulation of nuclear factor kappa B (NF-κB) p65 (P < 0.001), the protein content of tumor necrosis factor α (P = 0.033), and the mRNA abundance of IL-1β (P = 0.042) and nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3; P = 0.019). In contrast, pterostilbene inhibited the nuclear translocation of NF-κB p65 (P = 0.039) and suppressed the mRNA expression of IL-1β (P = 0.003) and NLRP3 (P = 0.049) in the jejunum. Moreover, pterostilbene administration induced a greater amount of reduced glutathione (P = 0.017) but a lower content of malondialdehyde (P = 0.023) in the jejunum of broilers compared with those received a basal diet. Overall, the current study indicates that dietary supplementation with pterostilbene may play a beneficial role in alleviating the intestinal damage of broiler chicks under the conditions of immunological stress.

Dataset Information

Effects of thymol and carvacrol supplementation on intestinal integrity and immune responses of broiler chickens challenged with Clostridium perfringens.

Publications

Effects of thymol and carvacrol supplementation on intestinal integrity and immune responses of broiler chickens challenged with Clostridium perfringens.

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