Dietary supplementation of prebiotic yeast Saccharomyces cerevisiae cell wall promotes growth performance and intestinal health in broiler chickens challenged with Clostridium perfringens.

1. This study evaluated the effectiveness of yeast (Saccharomyces cerevisiae) cell wall (YCW) supplementation on the growth performance, carcase characteristics, serum biomarkers, liver function, ileal histology and microbiota of broiler chickens challenged with Clostridium perfringens (C. perfringens).2. In a 35-d trial, 240 chicks aged 1-d-old were randomly assigned to one of four treatment groups, each with 10 replicates: control (CON) with no challenge or additives, challenged with C. perfringens (CHAL), CHAL and supplemented with YCW at either 0.25 g/kg (YCW0.25) or 0.5 g/kg (YCW0.5).3. In comparison to CON, the CHAL birds had reduced growth performance, survival rate, dressing percentage, breast meat yield, levels of total protein (TP), globulin (GLO), glucose (GLU), total antioxidant capacity (T-AOC) and total superoxide dismutase (T-SOD), as well as a decreased Lactobacillus population (P < 0.01). Additionally, this group showed elevated levels of glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), and C. perfringens count (P < 0.01). Compared to CHAL, the YCW0.25 or YCW0.5 groups had improved growth performance, survival rate, dressing percentage, breast meat yield, levels of TP, GLO, GLU, and T-AOC, as well as the activities of T-SOD, GOT, and GPT, villus height, villus surface area, villus height to crypt depth ratio, and the populations of both Lactobacillus and C. perfringens; (P < 0.01).4. The data suggested that YCW supplementation at either 0.25 or 0.50 g/kg can restore the growth performance of broiler chickens during a C. perfringens challenge.

Bromelain can reduce the negative effects of a subclinical necrotic enteritis in broiler chickens.

This study was conducted to examine the efficacy of a bromelain-based supplementation coded ANR-pf on growth performance and intestinal lesion of broiler chickens under necrotic enteritis (NE) challenge. A total of 540 Ross 308 day-old male chicks were randomly allocated into 6 treatments of 6 replicates. The bromelain formulation was delivered to chickens through gavaging or in drinking water method twice, on d 8 and 13. Nonchallenged groups included 1) without or 2) with the specific bromelain formulation gavaged at 0.8 mL/kg. NE-challenged groups included 3) without the specific bromelain formulation; 4) gavaged with 0.4 mL/kg; 5) gavaged with 0.8 mL/kg and 6) supplemented with 0.8 mL/kg via drinking water. Birds were challenged with Eimeria spp. on d 9 and Clostridium perfringens (NE-18 strain) on d 14 and 15. On d 14 and 19, fresh faecal contents were collected for the determination of oocyst counts. Intestinal lesion scores were determined on d16. Performance and mortality were recorded throughout the entire experiment. Among challenged groups, birds received additive via drinking water had higher weight gain (WG) compared to the remaining groups (P < 0.001) in the grower phase and had lower FCR compared to 0.4 mL/kg inoculated group in the grower and finisher phases (P < 0.001). Bromelain supplementation via drinking water improved the WG of challenged birds, similar to that of the nonchallenged birds (P < 0.001), and lowered FCR compared to other challenged groups (P < 0.001). Nonchallenged birds and birds that received bromelain formulation in drinking water did not have lesions throughout the small intestine whereas challenged birds, either un-supplemented or supplemented with bromelain via inoculation route recorded similar lesion score levels in the jejunum. At d 19, birds received bromelain in drinking water had lower fecal oocyst numbers compared to challenged birds without additive (P < 0.001). In conclusion, bromelain administration via drinking water could ameliorate the negative impacts of NE-infection in broilers by improving performance, lowering the oocyst numbers and lesion scores.

Effects of dietary Nisin on growth performance, immune function, and gut health of broilers challenged by Clostridium perfringens.

Nisin (Ni) is a polypeptide bacteriocin produced by lactic streptococci (probiotics) that can inhibit the majority of gram-positive bacteria, and improve the growth performance of broilers, and exert antioxidative and anti-inflammatory properties. The present study investigated the potential preventive effect of Nisin on necrotic enteritis induced by Clostridium perfringens (Cp) challenge. A total of 288 Arbor Acres broiler chickens of 1-d-olds were allocated using 2 × 2 factorial arrangement into four groups with six replicates (12 chickens per replicate), including: (1) control group (Con, basal diet), (2) Cp challenge group (Cp, basal diet + 1.0 × 108 CFU/mL Cp), (3) Ni group (Ni, basal diet + 100 mg/kg Ni), and (4) Ni + Cp group (Ni + Cp, basal diet + 100 mg/kg Ni + 1.0 × 108 CFU/mL Cp). The results showed that Cp challenge decreased the average daily gain (ADG) of days 15 to 21 (P<0.05) and increased interleukin-6 (IL-6) content in the serum (P < 0.05), as well as a significant reduction in villus height (VH) and the ratio of VH to crypt depth (VCR) (P<0.05) and a significant increase in crypt depth (CD) of jejunum (P<0.05). Furthermore, the mRNA expressions of Occludin and Claudin-1 were downregulated (P<0.05), while the mRNA expressions of Caspase3, Caspase9, Bax, and Bax/Bcl-2 were upregulated (P<0.05) in the jejunum. However, the inclusion of dietary Ni supplementation significantly improved body weight (BW) on days 21 and 28, ADG of days 15 to 21 (P<0.05), decreased CD in the jejunum, and reduced tumor necrosis factor-α (TNF-α) content in the serum (P<0.05). Ni addition upregulated the mRNA levels of Claudin-1 expression and downregulated the mRNA expression levels of Caspase9 in the jejunum (P<0.05). Moreover, Cp challenge and Ni altered the cecal microbiota composition, which manifested that Cp challenge decreased the relative abundance of phylum Fusobacteriota and increased Shannon index (P<0.05) and the trend of phylum Proteobacteria (0.05

Necrotic enteritis (NE), a severe digestive disorder in broiler chickens caused by Clostridium perfringens (Cp), a gram-positive bacterium, is a widespread issue in the global poultry industry, leading to significant economic losses. Nisin (Ni), a polypeptide bacteriocin produced by probiotic lactic streptococci, has been found to enhance daily weight gain and feed intake, while also exhibiting inhibitory effects on gram-positive bacteria and anti-inflammatory properties. In this study, a NE infection model in broilers was established to examine the potential preventive effects of Ni. These results demonstrated that Cp challenge reduced growth performance, caused inflammatory responses and intestinal apoptosis, damaged intestinal morphology and barrier function, and was accompanied by changes in the composition of the gut microbiota. Dietary supplementation with Ni improved growth performance and protected intestine against Cp challenge-induced damage in broilers. As a result, Ni may be a potential safe and effective additive for NE prevention in broiler production.

A microbial muramidase improves growth performance and reduces inflammatory cell infiltration in the intestine of broilers chickens under Eimeria and Clostridium perfringens challenge.

The objective of the present studies was to evaluate muramidase (MUR) supplementation in broilers under Eimeria and/or Clostridium perfringens challenge. For this, 2 experiments were conducted. Experiment 1. A total of 256 one-day old male Cobb 500 chicks were placed in battery cages in a completely randomized design, with 5 treatment groups, 7 replicate cages per treatment and 8 birds per cage. The treatments were: nonchallenged control (NC), challenged control (CC), CC + MUR at 25,000 or 35,000 LSU(F)/kg, and CC + Enramycin at 10 ppm (positive control-PC). Challenge consisted of 15× the recommended dose of coccidiosis vaccine at placement, and Clostridium perfringens (108 CFU/bird) inoculation at 10, 11, and 12 d. Macro and microscopic evaluation, immunohistochemistry, and gene expression were evaluated at 7, 14, 21, and 28 d of age. Experiment 2. A total of 1,120 one-day old male Cobb 500 chicks were placed in floor pens with fresh litter in a completely randomized design, with 4 treatment groups, 8 replicate pens per treatment, and 35 birds per pen. The treatments were: Control, supplementation of MUR at 25,000 or 45,000 LSU(F)/kg, and a positive control (basal diet plus Enramycin). At 10, 11, and 12 d of the experiment all the birds were inoculated by oral gavage with a fresh broth culture of a field isolate Clostridium perfringens (0.5 mL containing 106 CFU/bird). It was observed that in Experiment 1 MUR supplementation reduced the infiltration of macrophages and CD8+ lymphocytes in the liver and ileum of infected birds, downregulated IL-8 and upregulated IL-10 expression. In Experiment 2, MUR linearly improved the growth performance of the birds, increased breast meat yield, and improved absorption capacity. MUR supplementation elicited an anti-inflammatory response in birds undergoing a NE challenge model that may explain the improved growth performance of supplemented birds.

Drug-independent control strategy of clostridial infection in broiler chickens using anti-toxin environmentally friendly multienzymes.

The study investigated the effect of enzymes as a toxin detoxifier (DETOXIZYME) dietary supplementation on performance during growth, blood chemistry, and immunity under clostridia infection in chickens. A total of 480, day-old male chicks were randomly distributed to four groups, with six replicates of 20 birds each. The first control negative treatment (A) fed the basal formula as commercial feed prepared following the strain's needs, the second control positive group (B) fed the basal formula challenged with Clostridium perfringens (C. perfringens) type A, the third group (C) fed the basal formula with 100 g DETOXIZYME/ton of feed and challenged with clostridia, and the fourth group (D) fed the control basal formula with 100 g DETOXIZYME/ton of feed. DETOXIZYME dietary supplementation significantly boosted body weight (BW), body weight gain (BWG), feed intake (FI), and European production efficiency factor (EPEF) and improved the feed conversion rate (FCR) of the broilers. The dietary supplementation of DETOXIZYME significantly increased carcass trait and spleen. However, liver and abdominal fat weight significantly decreased compared with clostridia-challenged groups. The values of alanine aminotransferase (ALT), aspartate aminotransferase (AST), uric acid, creatinine, and Malondialdehyde (MDA) were decreased. While calcium, phosphate, zinc, and glutathione peroxidase (GPx) levels were improved in birds that took basal formulas fortified with DETOXIZYME contrary to the other treatment groups during 35 days of age. Plasma total cholesterol, triglyceride, and low-density lipoprotein (LDL) values were reduced versus the other treatment groups. Dietary supplementation of DETOXIZYME increased total protein, albumin, globulin, and Newcastle Disease (ND) immunity titer levels in the overall period compared to other groups. Dietary DETOXIZYME supplementation decreased clostridia and E. coli bacteria counts and improved gut morphometry. In conclusion, dietary supplementation of DETOXIZYME had a positive impact on performance, blood biochemistry, immunity, and bacterial counts and improved the gut morphology in broilers under clostridia infection.

Spray-dried porcine plasma enhances feed efficiency, intestinal integrity, and immune response of broilers challenged with necrotic enteritis.

Re-emergence of enteric diseases in the postantibiotic era has imposed severe loss to the poultry industry leading to the urgent need for appropriate additives to maintain gut health. Recently, more attention has been paid to animal plasma due to its high concentrations of active components such as albumins and globulins. The objective of this study was to evaluate the effects of spray-dried porcine plasma (SDP) supplementation during the starter phase (d 0-10) on growth performance, intestine health, and immune response of broilers under necrotic enteritis (NE) challenge. A total of 720 day-old male broiler parental line chicks (Ross 308) were randomly assigned to a 2 (NE challenge: no, yes) × 2 (SDP: 0, 2%) factorial arrangement with 12 replications of 15 chicks each. To induce NE, birds were inoculated with live Eimeria vaccine on d 9 and Clostridium perfringens on d 14. The body weight of birds and feed consumption were measured per pen on d 8, 10, 24, and 29 to calculate performance parameters. On d 16, three birds per pen were sampled to analyse the intestinal lesion score, gut permeability, villi morphology, relative weight of organs, and immune response. Results showed that SDP improved (P < 0.001) FCR in the pre-challenge phase (d 0-8). The results indicated that supplementing SDP lowered (P < 0.01) FCR at the end of the experiment (d 29). Dietary SDP decreased (P < 0.05) the concentration of FITC-d in serum samples of challenged broilers, although it did not affect the intestinal morphology and lesion score. Birds fed with SDP had a higher (P < 0.05) relative weight of bursa (g/kg live body weight) compared to non-supplemented birds. Supplementing SDP reduced the concentration of interleukin-6 (P < 0.05) and α-1 acid glycoprotein (P = 0.051) in serum samples of broilers. In conclusion, supplementation of SDP in the starter phase enhanced feed efficiency and gut integrity in NE challenged broilers, possibly through manipulating the immune response, while further studies targeting intestinal microflora and key genes are required to explore the mode of action.

Effects of dietary ß-1,3-glucan addition on the growth performance, mRNA expression in jejunal barrier, and cecal microflora of broilers challenged with Clostridium perfringens.

This experiment aimed to explore the interaction of ß-1,3-glucan and Clostridium perfringens on the growth performance, intestinal health and cecal microflora of broilers. A total of 384 one-day-old Arbor Acre broilers were sorted into 4 treatments with 6 replications. There were 2 factors in this trial: dietary ß-1,3-glucan addition including 0 and 250 mg/kg, intestinal enteritis challenged with Clostridium perfringens attack or not. Results showed that Clostridium perfringens infection disrupted the integrity of the intestinal mucosa by reducing the jejunal Occludin and Claudin-1 mRNA expression of broiler chickens at 21 d of age (P < 0.05). Meanwhile, when considering Clostridium perfringens as the main effect, it also decreased the mRNA expression of the glucose transporter recombinant sodium/glucose cotransporter 1 (SGLT1) at d 21 and the fatty acid transporter liver fatty acid-binding protein (L-FABP) at d 42 (P < 0.05) as well as affect cecum microbial diversity, especially in relative abundance of Firmicutes and Bacteroidetes. In addition, Clostridium perfringens infection reduced body weight, daily weight gain, and feed-gain ratio (FCR) in broilers at d 42 (P < 0.05). The dietary ß-1,3-glucan could alleviate intestinal mucosal damage caused by the Clostridium perfringens to some extent. When considering ß-1,3-glucan as the main effect, it increased the SGLT1 at 42 d of age (P < 0.05), and stabilized gut microbiota disorder caused by Clostridium perfringens. More over dietary ß-1,3-glucan addition increased body weight at 42-day-old (P < 0.05), and improved daily weight gain and FCR during 1 to 42 d (P < 0.05). In conclusion, dietary ß-1,3-glucan could improve growth performance and intestinal health in broilers infected with Clostridium perfringens.

A mixture of organic acids and thymol protects primary chicken intestinal epithelial cells from Clostridium perfringens infection in vitro.

Necrotic enteritis causes economic losses estimated to be up to 6 billion US dollars per year. Clinical and subclinical infections in poultry are also both correlated with decreased growth and feed efficiency. Moreover, in a context of increased antibiotic resistance, feed additives with enhanced antimicrobial properties are a useful and increasingly needed strategy. In this study, the protective effects of a blend of thymol and organic acids against the effects of Clostridium perfringens type A (CP) on chicken intestinal epithelial cells were investigated and compared to bacitracin, a widely used antibiotic in poultry production. Primary chicken intestinal epithelial cells were challenged with CP for a total time of 3 h to assess the beneficial effect of 2 doses of citric acid, dodecanoic acid, and thymol-containing blend, and compare them with bacitracin. During the challenge, different parameters were recorded, such as transepithelial electrical resistance, cell viability, mRNA expression, and reactive oxygen species production. CP induced inflammation with cytokine production and loss of epithelial barrier integrity. It was also able to induce reactive oxygen species production and increase the caspase expression leading to cellular death. The high dose of the blend acted similarly to bacitracin, preventing the disruptive effects of CP and inducing also an increase in zonula occludens-1 mRNA expression. The low dose only partially prevented the disruptive effects of CP but successfully reduced the associated inflammation. This study shows that the usage of thymol combined with 2 organic acids can protect primary chicken intestinal epithelial cells from CP-induced damages creating a valid candidate to substitute or adjuvate the antibiotic treatment against necrotic enteritis.

Clostridioides difficile infection and One Health: an equine perspective.

Clostridioides (Clostridium) difficile presents a significant health risk to humans and animals. The complexity of the bacterial-host interaction affecting pathogenesis and disease development creates an ongoing challenge for epidemiological studies, control strategies and prevention planning. The recent emergence of human disease caused by strains of C. difficile found in animals adds to mounting evidence that C. difficile infection (CDI) may be a zoonosis. In equine populations, C. difficile is a known cause of diarrhoea and gastrointestinal inflammation, with considerable mortality and morbidity. This has a significant impact on both the well-being of the animal and, in the case of performance and production animals, it may have an adverse economic impact on relevant industries. While C. difficile is regularly isolated from horses, many questions remain regarding the impact of asymptomatic carriage as well as optimization of diagnosis, testing and treatment. This review provides an overview of our understanding of equine CDI while also identifying knowledge gaps and the need for a holistic One Health approach to a complicated issue.

Mesoporous silica nanoparticles-based formulations of a chimeric proteinous vaccine candidate against necrotic enteritis disease.

To develop a nanoparticle-based vaccine against necrotic enteritis, a chimeric antigen (rNA) consisting of the main antigens of Clostridium perfringens, NetB, and Alpha toxin, was prepared. Then, the rNA molecules were loaded onto the functionalized mesoporous silica nanoparticles (MSNPs) using physical adsorption or covalent conjugation methods. The characterization of synthesized nanoparticles was performed by scanning electron microscopy, dynamic light scattering, zeta potential measurement, Fourier transform infrared spectroscopy, and thermogravimetry techniques. The results revealed that the spherical nanoparticles with an average diameter of 90 ±â€¯12 nm and suitable surface chemistries are prepared. MSNPs-based formulations did not show any significant toxicity on the chicken embryo fibroblast cells. The results of the challenge experiments using subcutaneous or oral administration of the as-prepared formulations in the animal model showed that the as-prepared nanosystems, similar to those formulated with a commercial adjuvant (Montanide), present stronger humoral immune responses as compared to that of the free proteins. It was also indicated that the best protection is obtained in groups vaccinated with MSNPs-based nanovaccine, especially those who orally received covalently conjugated nanovaccine candidates. These results recommend that the MSNPs-based formulated chimeric proteinous vaccine candidates can be considered as an effective immunizing system for the oral vaccination of poultry against gastrointestinal infectious diseases.

Buffered formic acid and a monoglyceride blend coordinately alleviate subclinical necrotic enteritis impact in broiler chickens.

The objective of this study was to evaluate the effect of 2 different doses of a partially buffered formic acid product (Amasil NA; 61% formic acid, 20.5% sodium formate), and a monoglyceride blend of short- and medium-chain fatty acids (BalanGut LS P) on necrotic enteritis (NE) infected broilers in terms of performance, intestinal microbial population and short-chain fatty acids concentrations in the gastrointestinal tract. A total of 528-day-old as hatched Ross 308 broilers were allocated to 48 pens with 11 birds in each pen. Six dietary treatments applied in the study were: T1) nonsupplemented diet (Control); T2) antibiotic supplemented diets; T3) and T4) high (Starter: 0.5%; Grower and Finisher: 0.5%) and low (Starter: 0.3%; Grower and Finisher: 0.2%) dose of Amasil NA; and groups T5) and T6) high (Starter: 0.3%; Grower and Finisher: 0.2%) and low dose (Starter: 0.3%; Grower: 0.15%; Finisher: 0.075%) of (BalanGut LS P). All birds in this study were fed starter (d 0-10), grower (d 11-24) and finisher (d 25-35) diets and challenged with NE. To induce subclinical NE, oral administrations of Eimeria oocysts (d 9) followed by inoculation of Clostridium perfringens strains (d 14 and 15) were applied. Results showed that birds fed the high dose of Amasil NA, had a higher feed conversion ratio (FCR,P < 0.05) compared to the nonsupplemented group during the starter period. Antibiotic supplementation reduced FCR during the grower (P < 0.001), finisher (P < 0.05) and overall (P < 0.001) periods of the experiment. Both levels of BalanGut LS P and low levels of Amasil NA enhanced overall FCR (P < 0.05) compared to the birds in the nonsupplemented group. Compared to the nonsupplemented group, high levels of Amasil NA and low levels of BalanGut LS P improved FCR in the finisher stage (P < 0.05). On d 16, cecum digesta of birds fed with antibiotic supplemented diets showed a significantly lower number of C. perfringens (P < 0.001) compared to the nonsupplemented and high level of BalanGut LS P group. Bacillus (P < 0.01) and Ruminococcus numbers were significantly lower in the birds fed with high level of Amasil NA (P < 0.05) compared to the antibiotic supplemented diets. High doses of Amasil NA, showed the highest propionate concentration in the cecum (P < 0.001). The study suggests that supplementation of BalanGut LS P and Amasil NA at different feeding phases may achieve optimal performance improvement in broilers under NE challenge.

Evaluation of the therapeutic effect of propolis on Fasciola gigantica and Clostridium novyi type B infections in sheep.

The use of natural products for disease control is a promising approach to solving the problem of drug resistance. The aim of the research reported here was to evaluate the fasciolicidal and anti-Clostridium novyi type B activities of propolis administered orally to sheep infected with Fasciola gigantica and C. novyi type B. Sheep infected with both pathogens were divided into two groups: an infected treated group and an infected non-treated group. The treatment was oral administration of 50 mg propolis extract/kg daily for 15 days. The body weight of the sheep, fecal egg counts of F. gigantica, serum levels of F. gigantica IgG, concentrations of cytokines (IL-2, IL-10, and IL-17), and bacterial counts of C. novyi were evaluated. Following treatment, the sheep had increased body weight and a significant decrease in the egg count, which was reduced by 54.54% at 15 days post treatment. The level of anti- Fasciola IgG increased, whereas levels of IL-2, IL-10, and IL-17 decreased in propolistreated sheep. Treatment of sheep with propolis produced a significant reduction in fecal count of C. novyi, from 8 × 109 to 3 × 103 colony units per gram at 15 days post treatment. This research highlights the therapeutic potential of Egyptian propolis extract as a treatment against F. gigantica and C. novyi type B infections, and investigated its mode of action through its effect on some cellular and humoral responses in sheep with both infections.

Immunomodulatory effect of short-term supplementation with Bacillus toyonensis BCT-7112T and Saccharomyces boulardii CNCM I-745 in sheep vaccinated with Clostridium chauvoei.

The bacterium Clostridium chauvoei is the causative agent of blackleg in livestock, and vaccination is the most effective means of prevention. The aim of this study was to assess the effect of short-term supplementation with Bacillus toyonensis and Saccharomyces boulardii on the immune response to a C. chauvoei vaccine in sheep. Sheep were vaccinated subcutaneously on day 0 and received a booster dose on day 21, with 2 mL of a commercial vaccine formulated with inactivated C. chauvoei bacterin adsorbed on aluminum hydroxide. Probiotics were orally administered B. toyonensis (3 × 108 cfu) and S. boulardii (3 × 108 cfu) over five days prior to the first and second doses of the vaccine. Sheep supplemented with B. toyonensis and S. boulardii showed significantly higher specific IgG, IgG1, and IgG2 titers (P<0.05), with approximately 24- and 14-fold increases in total IgG levels, respectively, than the nonsupplemented group. Peripheral blood mononuclear cells from the supplemented group had increased mRNA transcription levels of the IFN-γ, IL2, and Bcl6 genes. These results demonstrate an adjuvant effect of short-term supplementation with B. toyonensis and S. boulardii on the immune response against the C. chauvoei vaccine in sheep.

Effects of feed supplementation with 3 different probiotic Bacillus strains and their combination on the performance of broiler chickens challenged with Clostridium perfringens.

The application of probiotics in broiler feed, to alleviate performance deficiencies due to mild infections by coccidia and Clostridium perfringens, is of increasing interest for the poultry industry. Therefore, our objective was to evaluate the capacity of 3 Bacillus strains and their combination as probiotics in vitro and in vivo. Thus, protein and carbohydrate degradation and C. perfringens growth inhibition capabilities were assessed by colometry measurement and an agar diffusion bioassay, respectively. A total of 2,250 1-day-old male broiler chicks were assigned to 5 dietary treatments: 1) non-probiotic-supplemented control (control); 2) control + DSM 32324 at 0.8 × 106 cfu/g of feed; 3) control + DSM 32325 at 0.5 × 106 cfu/g of feed; 4) control + DSM 25840 at 0.3 × 106 cfu/g of feed; and 5) control + DSM 32324 + DSM 32325 + DSM 25840 at 1.6 × 106 cfu/g of feed. A pathogenic field strain of C. perfringens was used to induce the necrotic enteritis challenge on day 19, 20, and 21. All birds and remaining feed were weighed on pen basis on day 0, 21, 35, and 42, to calculate BW gain and mortality-adjusted feed conversion. Mortality and mortality due to necrotic enteritis were recorded daily. On day 21, 45 birds per treatment were evaluated for macroscopic intestinal necrotic enteritis lesions. Performance data were statistically analyzed using an ANOVA and subjected to a least significant difference comparison. Necrotic enteritis lesion scores were statistically analyzed using nonparametric Kruskal-Wallis test. Dunn's test was used for treatment comparison. The tested strains showed different abilities of degrading protein and carbohydrates and inhibiting C. perfringens growth in vitro. The birds fed the multi-train combination presented significantly better performance and lower necrotic enteritis lesion score than those in the control group. Dietary supplementation with probiotics resulted in significantly lower necrotic enteritis mortality. The results demonstrate the suitability of the evaluated Bacillus multistrain combination as an effective probiotic in C. perfringens-challenged chickens.

Study on the interactive effect of deoxynivalenol and Clostridium perfringens on the jejunal health of broiler chickens.

With global warming and ban on antibiotics, it occurs occasionally that deoxynivalenol (DON) together with Clostridium perfringens impairs the gut health of broiler chickens. However, the interactive effect of DON and C. perfringens on intestinal health is still unknown. A total of 120 one-day-old Arbor Acres broilers were randomly distributed to 4 groups. Birds were gavaged with C. perfringens (8 × 108 CFU/d per bird) or sterile medium and fed a DON diet (0 or 5 mg of DON per kg diet) to investigate the interactive effects. The main effect analysis showed that DON diet significantly downregulated (P < 0.05) the mRNA expression of mucin-2, B-cell lymphoma-2-associated X, and cysteinyl aspartate-specific proteinase-3 of jejunal mucosa; decreased (P < 0.05) the indexes of ACE, Chao1, Shannon, and Simpson; and also decreased the relative abundance of the phylum Bacteroidete and the genera Lactococcus in jejunal contents of broilers chickens. Meanwhile, C. perfringens significantly increased (P < 0.05) crypt depth; decreased (P < 0.05) the ratio of villi height to crypt depth, the activity of jejunal diamine oxidase, and the relative abundance of Lactococcus; and upregulated (P < 0.05) the relative expression of B-cell lymphoma-2 and cysteinyl aspartate-specific proteinase-8. Furthermore, the interactions between DON and C. perfringens were most significant (P < 0.05) in the mRNA expression of lipopolysaccharide-induced TNF factor (LITAF) and TLR-4, the abundance of the genera Lactococcus in jejunal contents, and butyric acid concentrations in cecal contents of birds. Finally, Spearman correlation analysis suggested that the most negative correlations (P < 0.05) with the abundance of the genera except Lactobacillus were observed within the mRNA expression of LITAF. The abundance of Lactococcus had a positive correlation (P < 0.05) with the expression of Caspase-3. Most genera except Lactobacillus negatively correlated (P < 0.05) with acetic acid, butyric acid, and total short-chain fatty acids. In conclusion, dietary deoxynivalenol and C. perfringens challenge had a harmful effect on the jejunal health and should be carefully monitored in broiler production.

Varying starch to fat ratios in pelleted diets: II. Effects on intestinal histomorphometry, Clostridium perfringens and short-chain fatty acids in Eimeria-challenged broiler chickens.

1. The hypothesis behind the study was that a high dietary starch level (HS) would lead to impaired gut health compared to a low-starch diet (LS) in Eimeria-challenged broilers. The effects of two diets with different starch to fat ratios on intestinal histomorphometry, Clostridium perfringens counts and toxin profile, necrotic enteritis prevalence and abundance of short-chain fatty acids (SCFAs) were examined. 2. A total of 1,920 one-day-old Ross 308 broiler chickens were fed one of two isocaloric diets formulated either with high (32:1) or low (2:1) starch to fat ratios from d 10 to 29 of age. Each treatment group had 12 pen replicates containing 80 broilers each. On d 17, the chickens were challenged with Eimeria vaccine strains. Samples were collected on d 16, 21-23 and 29. 3. Whereas villus length increased gradually throughout the study in the HS group, a peak level was reached on d 21-23 in the LS group. On d 29, the HS group had significantly longer villi than the LS group. 4. Caecal SCFA concentrations were higher in the HS group compared to the LS group on d 16. In both groups, the SCFA level peaked on d 21-23, with the most pronounced increase seen in the LS group. 5. The C. perfringens netB:cpa ratio increased from d 16 to 29 in the HS group. C. perfringens counts and necrotic enteritis prevalence were similar between the two groups. 6. Diet affected the dynamics of small intestinal villus length and caecal SCFA abundance. These findings suggest that structural remodelling of the small intestine is an adaptation to different dietary starch levels, and that caecal SCFA abundance is associated with the availability of substrate for the microbiota in the posterior intestinal segments. Chickens adapted to higher levels of dietary starch might be more robust against Eimeria infections due to increased mucosal surface area. Studies with other dietary starch sources are required to clarify the impact of dietary starch levels on intestinal health in Eimeria-challenged broilers.

Cannabis-derived cannabidiol and nanoselenium improve gut barrier function and affect bacterial enzyme activity in chickens subjected to C. perfringens challenge.

Revealing the multifocal mechanisms affecting cross-talk between Clostridium perfringens pathogenesis and the host response is an urgent need in the poultry industry. Herein, the activity of Cannabis sativa-derived cannabidiol (CBD) and selenium nanoparticles (Nano-Se) in modulating the host response to Clostridium perfringens challenge was investigated in broiler chickens subjected to a mild infection model. The infected chickens exhibited no clinical manifestations, confirming the potential hazard of pathogen transmission to the food chain in the commercial sector. However, both CBD and Nano-Se affected the responses of chickens to C. perfringens challenge. The beneficial actions of both agents were manifested in the upregulated expression of genes determining gut barrier function. Both CBD and Nano-Se promoted shifts in gut bacterial enzyme activity to increased energy uptake in challenged chickens and upregulated potential collagenase activity. There was no opposite effect of CBD and Nano-Se in mediating the host response to challenge, whereas an additive effect was evidenced on the upregulation of gene determining gut integrity. Collectively, these findings indicate that understanding the action mechanisms of CBD and Nano-Se is of great interest for developing a preventive strategy for C. perfringens infection in broilers.

Efficacy of Trace Mineral-Amino Acid Complexes in a Necrotic Enteritis Challenge Model.

Necrotic enteritis (NE) is a common and costly disease of poultry caused by virulent toxigenic strains of Clostridium perfringens. Although the importance of trace minerals for intestinal integrity and health is well documented, there is little information on their role in ameliorating the effects of NE. The two studies reported here examined the effects of replacing a portion of the dietary zinc (Zn), copper (Cu), and manganese (Mn) supplied as sulfates in the control diets with metal-amino acid-complexed minerals in a NE-challenge model consisting of coccidiosis and Clostridium perfringens. In a 28-day battery study, the treatments were the following: (1) no additional Zn or Mn, unchallenged (negative control); (2) no added Zn or Mn, challenged (positive control); (3) added ZnSO4 and MnSO4 at 100 ppm each, challenged; (4) additional ZnSO4 at 60 ppm, Availa-Zn at 40 ppm (Low), and MnSO4 at 100 ppm, challenged; (5) added ZnSO4 at 60 ppm, Availa-Zn at 60 ppm (high), and MnSO4 at 100 ppm, challenged; and (6) added ZnSO4 at 60 ppm, Availa-Zn at 40 ppm, MnSO4 at 60 ppm, and Availa-Mn at 40 ppm, challenged. None of the treatments ameliorated gross lesion scores, but all reduced NE-associated mortality compared with the positive control. At 28 days, the group supplemented with Availa-Zn at 40 ppm (low) had a lower body weight than challenged groups supplemented with Zn and the negative control. In a floor pen study, the five treatment groups were the following: (1) Zn, Mn, and Cu from sulfate sources at 100, 100, and 20 ppm respectively; (2) Zn, Mn, and Cu from sulfate sources at 40, 100, and 20 ppm, respectively, plus Zn from Availa-Zn at 60 ppm; (3) Zn and Mn from sulfate sources at 40 and 100 ppm, respectively, plus Zn from Availa-Zn at 60 ppm and Cu from Availa-Cu at 10 ppm; (4) Zn, Mn, and Cu from sulfate sources at 60, 60, and 20 ppm, respectively, plus Zn and Mn from Availa-Zn/Mn at 40 and 40 ppm, respectively; and (5) bacitracin methylene disalicylate at 55 g/metric ton with Zn, Mn, and Cu from sulfate sources at 100, 100, and 20 ppm, respectively (Zoetis, Inc., Kalamazoo, MI). None of the treatments reduced lesion scores. The Availa-Zn and Availa-Zn/Mn had lower mortality than the sulfate-supplemented feed, whereas Availa-Zn/Cu and bacitracin methylene disalicylate were intermediate and did not differ from the other groups. Considering both trials together, and by using NE mortality as the discriminating factor, we found that adding Zn and Mn exceeding National Research Council requirements reduced NE-associated mortality, and in the floor pen study, complexed Zn and complexed Zn plus Mn appeared to be superior to sulfates.

Effects of Replacing In-feed Antibiotics with Synergistic Organic Acids on Growth Performance, Health, Carcass, and Immune and Oxidative Statuses of Broiler Chickens Under Clostridium perfringens Type A Challenge.

This study was conducted to investigate the effects of replacing in-feed antibiotics with synergistic organic acids on growth performance, health, carcass, and immune and oxidative statuses of broiler chickens under Clostridium perfringens (CP) type A challenge. Two organic acid products were tested: organic acid 1 (OA1), consisting of butyrate, medium-chain fatty acids, organic acids, and phenolics; and organic acid 2 (OA2), consisting of buffered short-chain fatty acids. Six hundred 1-day-old male Arbor Acres broiler chicks were randomly assigned to one of five treatments: Control 1, basal diet, nonchallenged birds; Control 2, basal diet, with CP challenge; antimicrobial growth promoters (AGP), basal diet supplemented with Aureomycin (chlortetracycline), with CP challenge; OA1, basal diet supplemented with OA1, with CP challenge; and OA1OA2, basal diet supplemented with OA1 and OA2, with CP challenge. Each treatment had eight replicate pens of 15 birds. The experiments lasted for 29 days. The disease challenge was performed on days 15-17, with an oral gavage of 0.5 mL of CP culture (2.0 × 108 colony-forming units [CFU]/mL) for each bird. Body weights (BWs), intestinal lesion scores, immune organ indices, and serum malondialdehyde (MDA) concentrations were measured on days 19, 22, and 29, respectively, in three birds per pen. Carcass characteristics were determined on day 29. No treatment-related differences in mortality were noted before (P = 0.28) or after (P = 0.64) challenge or over the whole study period (days 0-28; P = 0.66). On day 19, the BW of Control 2 was lower than other treatments (P < 0.0001). On day 22, AGP, OA1, and OA1OA2 had higher BW than Control 2 (P = 0.001). The breast muscle yield of OA1 and OA1OA2 was higher than AGP (P < 0.05). The abdominal fat yield of OA1OA2 was lower than AGP and Control 2 (P < 0.05). On day 22, the birds fed OA1OA2 showed lower intestinal lesion scores than OA1 (P < 0.05). No treatment-related differences in immune organ (spleen, thymus, and bursa) indices were noted (P > 0.05). On day 29, the MDA concentration of OA1 and OA1OA2 was lower than those of Control 1 and AGP (P < 0.05). In conclusion, the addition of organic acids may protect broiler chickens from severe intestinal lesions and oxidative stress and may help reduce abdominal fat mass deposition. There is potential for organic acid-based products as alternatives for AGP in preventing necrotic enteritis in broilers.

Effect of Bacillus subtilis DSM 32315 under Different Necrotic Enteritis Models in Broiler Chickens: A Meta-Analysis of Five Independent Research Trials.

Challenge models are needed to understand the pathogenesis of necrotic enteritis (NE) and provide the basis of evaluating nonantibiotic feed-additive interventions. In the category of nonantibiotic feed additives, the application of probiotics to improve intestinal health and growth performance of broiler chickens in the face of an NE challenge has been well described. However, it is crucial to evaluate the consistency of specific probiotics for mitigating the disease challenge and improving performance. Therefore, a meta-analysis of five independent research trials was conducted with the objective of evaluating the effect of Bacillus subtilis DSM 32315 (probiotic) on body weight gain (BWG), feed conversion ratio (FCR), NE mortality, and lesion score (LS) of broiler chickens challenged with NE. These independent studies were conducted in three countries (the United States, Thailand, and Finland). The statistical analysis used fixed and random effects to estimate the mean effect size (MES) of the difference between NE-challenged birds (control) and NE-challenged probiotic-fed birds and the 95% confidence interval of MES. A meta-regression was performed to evaluate heterogeneity (MES variance) among studies. The statistical analysis was performed using a robust variance estimation strategy with a SAS macro. Probiotic-supplemented birds had a significantly higher BWG (MES = 1.04, P = 0.009) and a significantly lower FCR (MES = -1.39, P = 0.020), NE mortality (MES = -1.15, P = 0.012), and LS (MES = -1.29, P = 0.045). Response variables of BWG (Q = 2.81, P = 0.560) and NE mortality (Q = 5.60, P = 0.354) did not present heterogeneity. Heterogeneity was found for FCR (Q = 10.34, P = 0.035) and LS (Q = 16.13, P = 0.001). Overall, dietary supplementation of B. subtilis DSM 32315 significantly improved BWG and reduced FCR, mortality, and LS in a repeatable large-scale manner.