Symposium on Gut Health in Production of Food Animals 2021

Glenn Zhang, Oklahoma State University

The rising concern of antimicrobial resistance highlights a need for effective alternatives to antibiotics for livestock production. Butyrate, forskolin, and lactose are 3 natural products known to induce the synthesis of host defense peptides (HDP), a critical component of innate immunity. In this study, the synergy among butyrate, forskolin, and lactose in enhancing innate host defense, barrier function, and resistance to necrotic enteritis and coccidiosis was investigated. Our results indicated that the 3 compounds synergistically augmented the expressions of multiple HDP and barrier function genes in chicken HD11 macrophages. The compounds also showed an obvious synergy in promoting HDP gene expressions in chicken jejunal explants. Dietary supplementation of a combination of 1 g/kg sodium butyrate, 10 mg/kg forskolin-containing plant extract, and 10 g/kg lactose dramatically improved the survival of chickens from 39% to 94% (P < 0.001) in a co-infection model of necrotic enteritis. Furthermore, the 3 compounds largely reversed growth suppression, significantly alleviated intestinal lesions, and reduced colonization of Clostridium perfringens or Eimeria maxima in chickens with necrotic enteritis and coccidiosis (P < 0.01). Collectively, dietary supplementation of butyrate, forskolin, and lactose is a promising antibiotic alternative approach to disease control and prevention for poultry and possibly other livestock species.

Key Words: antibiotic alternative, host defense peptide, butyrate, necrotic enteritis, coccidiosis.

Authors: Q. Yang and G. Zhang*, Oklahoma State University, Stillwater, OK, USA

Hongyu Xue, Amlan International

Homeostasis of the gut microbial ecosystem is essential for optimal growth performance and disease resistance in postweaning pigs. A220 is a formulated feed additive that features a blend of a proprietary toxin-adsorbing mineral with a select blend of phytogenics shown to have antibacterial properties against a variety of gram-negative and positive bacterial pathogens. This study aimed to assess the effects of A220 on the gut microbiome and further relate this to changes in growth performance and post-weaning diarrhea outcomes in pigs challenged with an enterotoxigenic Escherichia coli (ETEC). At 21 d of age, 36 piglets were weaned and randomly allocated to 1 of 3 groups: control (CON) or A220 supplemented at 0.25 or 0.5%. After a 7-d adaptation, all pigs were orally inoculated with 1010 cfu of F18 ETEC once daily from d 0 to d 2 post-inoculation (PI). Fecal consistency was scored twice daily from d 0–21 PI. Microflora in ileal digesta, ileal mucosa and fecal samples were profiled using 16S rRNA sequencing on d −7, 0, 7, and 21 PI. A220 supplementation at both levels increased feed efficiency from d 14–21 PI and reduced diarrhea frequency during the study (P < 0.05). Compared with CON, 0.5% A220 supplementation increased the relative abundance of Firmicutes, but reduced Bacteroidetes and Proteobacteria in feces on d 7 PI (P < 0.05). Within Firmicutes phylum, A220 at both 0.25% and 0.50% increased the relative abundance of Lactobacillaceae on d 7 PI (P < 0.05). A220 at both levels increased the relative abundance of Prevotellaceae and decreased that of Rikenellaceae in ileal digesta on d 21 PI (P < 0.05). A220 at 0.5% increased the relative abundance of Clostridiaceae and Prevotellaceae, and decreased that of Enterobacteriaceae and Succinivibrionaceae in ileal mucosa on d 21 PI (P < 0.05). A220 supplementation modified gut microbiota in favor of promoting a well-balanced gut microbial ecosystem, which may contribute to enhanced disease resistance and improved growth performance in weanling pigs faced with pathogenic challenges.

Key Words: microbiome, enterotoxigenic Escherichia coli (ETEC), post-weaning, pig.

Authors: H. Xue* 1, D. Wang 1, L. Johnston 1, Y. He 2, C. Jinno 2, Y. Liu 2, and P. Ji 3 / 1 Amlan International, Chicago, IL, USA, 2 Department of Animal Science, University of California, Davis, CA, USA, 3 Department of Nutrition, University of California, Davis, CA, USA.

Jane Leedle, JL Microbiology

Antibiotics and zinc oxide have long been used to prevent or cure post-weaning diarrhea. Now many countries are reducing use or completely banning these substances due to antimicrobial resistance and environmental concerns. Without antimicrobials producers have increased demand for natural solutions. Lactobacilli postbiotics are known to have a strong effect in reducing diarrhea in children. Research suggests this postbiotic prevents pathogen adherence and shifts the gut microbiome toward beneficial flora. Our objective was to evaluate the postbiotic Lactobacillus LB on newly weaned piglet health and performance outcomes compared with Control and Zinc oxide (Zn) in a 42-d E. coli challenge study. Three hundred male pigs 18 to 22 d of age and 2.75 to 7.60 kg BW from a commercial farm were divided into 4 groups each having 15 pens of 5 piglets. The LB treatment was Control feed with 2 kg LB/MT. The Zn treatment was Control with 3000 ppm Zn fed for 21 d and then removed. The 4th treatment was LB+Zn: Control diet with LB and added Zn for the 1st 21 d then Zn was removed. On d 10, all pigs were gavaged with 5 mL of 1.3 × 10e9 cfu E. coli/mL. Response variables were clinical and fecal scores, ADG, and FCR, with data collected on specific study days. Statistical analyses were performed using SAS and a Tukey-Kramer test. Fecal score results showed a strong reduction in diarrhea in the LB group. No watery stool was present in any pen in the LB group, yet it was as high as 89% in Control and 7% in the Zn group. Duration was 5 d post-challenge in LB as opposed to 11 and 7 d for Control and Zn groups, respectively. LB+Zn reduced diarrhea to 5 d as well. The E. coli challenge was severe. Mortality ~40% in the Control group was and 25% in the LB group. At 42 d postweaning, LB piglets had better growth performance. FCR was 13% lower (1.00 vs 1.15; P < 0.01) and ADG was 50% higher (0.21 kg/d vs 0.14 kg/d; P < 0.01) compared with Control. ADG was not different between LB and Zn groups. In summary, the postbiotic Lactobacillus LB significantly reduced postweaning diarrhea and was a good alternative to zinc oxide in piglet production.

Key Words: piglet, postweaning diarrhea, postbiotic.

Authors: P. Tacon 1, S. Della Zassa 2, C. Cull 3, K. Lechtenberg 3, and J. Leedle *4 / 1 ADARE Biome, Houdan, France, 2 Adare Biome, Lawrenceville, NJ, USA, 3 Midwest Veterinary Services Inc., Oakland, NE, USA, 4 JL Microbiology, Hartland, WI, USA.

Luis-Miguel Gomez-Osorio, Alura Inc.

The aim of this work was to determine the in vitro effectiveness of a blend of short-chain fatty acids (SCFA) and medium-chain fatty acids (MCFA) using α-monoglycerides through a minimum inhibitory concentration (MIC) test. Clostridium perfringens type G strains used in this study were previously isolated from field cases of focal duodenal necrosis (FDN) and necrotic enteritis (NE) (Villegas et al., 2020, J. Vet. Diagn. Invest., 32:268–276). Isolated colonies of the tested bacterial species were selected from a 24-h culture on a blood agar plate. MIC tests were done in triplicates using 2 samples of Fractal and 3 strains of Clostridium perfringens. The strains were isolate 1 and isolate 5 (obtained from chickens with FDN) and CP6 (obtained from a broiler chicken with NE). The microbicidal activity of the SCFA and MCFA α-monoglyceride product for CP6 strain is shown in Figures 1, 2 and 3. All isolates had MICs ≤0.08 μg/mL. Furthermore, the MICs for isolate 1 and isolate 5 were 0.04 respectively. For CP6 strain, the MIC was 0.08. The blend of SCFA and MCFA α-monoglycerides demonstrated high antimicrobial activity against different strains of pathogenic Clostridium perfringens isolated from field cases of FDN and NE under an in vitro test.

Key Words: focal duodenal necrosis, necrotic enteritis, α-monoglycerides, short-chain fatty acids, medium-chain fatty acids.

Authors: L.-M. Gomez-Osorio *1, L. Stabler 2, and M. Franca 2 / 1 Alura Inc., Durham, NC, USA, 2 Department of Population Health, Poultry Diagnostic and Research Center, University of Georgia, Athens, GA, USA.

Guillermo Tellez-Isaias, University of Arkansas

Several phytogenics have been evaluated as feed additives in the poultry industry for nutritional purposes. However, phytogenics play an essential role in the prevention of several diseases in poultry due to their antioxidant, anti-inflammatory, antibacterial, antiviral, antifungal, and immunomodulatory properties. Hence, in recent years, our laboratories have studied several phytogenics as feed additives to control salmonellosis and necrotic enteritis, to reduce the severity of aflatoxicosis in broiler chickens, and to control coccidiosis in Leghorn chickens. Curcumin is a bright yellow chemical and the principal curcuminoid of turmeric (Curcuma longa), a member of the ginger family (Zingiberaceae). The aim of this presentation is to summarize the studies that have been published by our laboratories evaluating this remarkable phytochemical alone and in combination with other nutraceuticals: (1) Hernandez et al., 2018, Front. Microbiol. 9:1289; (2) Hernandez et al., 2019, Animals 9:184; (3) Solis et al., 2019, Toxins 11:121; (4) Leyva et al., 2021, J. Anim. Sci. Biotechnol. 12:23; (5) Petrone et al., 2021, Sci. Rep. 11:1–9.

Key Words: curcumin, salmonellosis, coccidiosis, aflatoxicosis, necrotic enteritis.

Authors: G. Tellez-Isaias *1, D. Hernandez-Patlan 2, B. Solis-Cruz 2, V. Petrone-Garcia 2, A. Leyva-Diaz 2, C. Vuong 1, D. Graham 1, C. Selby 1, J. Latorre 1, and B. Hargis 1 / 1 University of Arkansas, Fayetteville, AR, USA, 2 UNAM, Cuautitlan Cuautilan Izcalli, Mexico, Mexico.

Don Ritter, Innovad USA

Compromised intestinal health in modern production has been linked with chronic inflammation and significant economic losses. Although several challenges are artificially introduced in recent poultry models of gut failure/dysbiosis, the ‘too clean’ experimental conditions oppose a major limitation. Here we evaluated (1) a model of chronic gut inflammation under real farming conditions using, as a trigger, high non-starch polysaccharides (without NSPases), and (2) the ability of an in-feed technology (Lumance Innovad: esterified butyrate with plant extracts and essential oils) to alleviate inflammatory responses in broilers. In experiment 1, the high NSP (60% wheat + 5% rye) significantly increased macroscopic morphometric “dysbacteriosis” both at d 21 and d 28 over a standard diet (50% corn + 30% soy) (P < 0.005; 1 bird per pen scored, n = 8 pens/group; n = 30 birds/pen), evaluated according to Teirlynck et al. (2011). In experiment 2, a heatwave elevated the mean temperature to ~32°C for ~12–14 h/d inside the production between d 21 and d 29. Interestingly, although both intestinal and systemic oxidative stress (measured as MDA) in the high NSP diet reduced by ~30% between d 28 and d 35, both inflammatory (IFN-γ) and immune (IgA) responses increased significantly (~33 and 40%, respectively) suggesting a cumulative impact over time. Importantly, Lumance (1 and 2 kg/ton) significantly increased the BW at the end of the life cycle (d 35) over the high-NSP (P= 0.041) and reduced the FCR (P = 0.012) (n = 8 pens/group; n = 30 birds/pen). Also, Lumance reduced, in a dose response manner, both intestinal and systemic MDA at d 28 (P < 0.05), intestinal IFN-γ at d 28 and d 35 (P < 0.05) and numerically both intestinal and systemic IgA at d 28 and d 35. In conclusion, we have successfully established a novel, dietary-induced model of chronic gut inflammation under real farming conditions, which can be seen as the sum of several stress points and evaluated the ability of an in-feed technology (Lumance Innovad) to alleviate its inflammatory responses in broilers.

Key Words: gut health, chronic inflammation model, real farming condition, biomarker, feed additive.

Authors: A. Khadem 1,2, D. Ritter *3 , and C. Gougoulias 1 / 1 Innovad NV/SA, Essen, Belgium, 2 Lab of Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium, 3 Innovad USA, Salisbury, MD, USA.

Audrey Duff, The Ohio State University

Heterophil granule component β-d-glucuronidase has been used to assess degranulation activity in cell culture supernatant after bacterial challenge. Adaptation of this assay for site specific degranulation in gastrointestinal (GIT) tissue was evaluated as a potential indicator of localized inflammation in response to Eimeria maxima (EM) infection. Experiments (Exp) 1–3 compared Control (C1, 2, 3) and EM-infected groups (EM1, 2, 3) by t-test (P < 0.05), while Experiment 4 used ANOVA and Dunnett’s post hoc analysis (P < 0.05) to compare C4 to EMlow (5 × 10³ oocysts), -medium (1 × 10⁴ oocysts), and –high (1.5 × 10⁴ oocysts). Intestinal scrapings were collected into RPMI 1% penicillin/streptomycin on ice, gently homogenized, and centrifuged. Supernatants were incubated with RPMI and positive control with serum opsonized zymosan for 1 h at 42°C. Samples were tested in 3 replicates and incubated with 4-methylumbelliferyl-β-d-glucuronide for 4 h at 42°C. Liberated 4-methylumbelliferone (4-MU) was quantified fluorometrically at an excitation/emission wavelength of 360/460 nm. Body weight gain (BWG) was evaluated over the infection period and was significantly suppressed in all EM groups relative to C1–3 (P < 0.05), while only EM-high BWG was lower (P = 0.008) than C4. In Exp 1 and 2, EM1 and EM2 4-MU was significantly lower than C1 and C2 (P = 0.046 and 0.009, respectively), while no differences were observed between C3 and EM3 in Exp 3 (P = 0.351). Exp 4 showed significant decrease in EM-low (P = 0.003) and EM-high (P = 0.003) 4-MU versus C4, but not EM-medium (P = 0.357). All assay positive controls were elevated relative to samples. Suppression of BWG indicates successful EM infection and supports the conclusion of a general observable decease in degranulation in the GIT of EM-challenged broilers. In future iterations, addition of correlation analyses between 4-MU values, BWG, and EM lesion scores on a per bird basis would aid in addressing variability within treatment groups. These results may provide new insight into GIT degranulation events occurring during coccidial infection.

Key Words: degranulation, Eimeria, gastrointestinal (GIT), inflammation, broiler.

Authors: A. Duff*, K. McGovern, M. Trombetta, and L. Bielke, The Ohio State University, Columbus, OH, USA.

D. Ayala, Land O' Lakes

Cellulitis in commercial turkeys has emerged as one of the leading causes of morbidity and mortality in the United States. According to the United States Animal Health Association industry survey, cellulitis ranks among the top 5 health issues for the turkey industry. It also represents a major cause of carcass condemnation at slaughter with significant economic losses for turkey producers. The main bacterial pathogens associated with the disease are Clostridium spp., avian pathogenic Escherichia coli, and Staphylococcus aureus. The aim of this study was to characterize the microbial community of birds with cellulitis and compare it to the microbial profile of healthy birds, to identify potential agents and routes of the infection. A total of 4 sample types including cecum, ileum, skin, and subcutaneous tissue (SBT) from 10 Nicholas turkeys (1) with cellulitis (Cell+ group), (2) without cellulitis (Cell− group), and (3) healthy birds (Control group) were collected between 16 and 18 weeks of age. Samples of Cell+ and Cell− groups were collected at one farm with a history of cellulitis, whereas samples of Control group were collected at a sister farm, with no history of cellulitis. The microbial profile of all samples was characterized by 16S metagenomics. The SBT microbiome of Cell+ samples was dominated by Clostridium sensu stricto with a relative abundance of 65.86% compared with 0.06% and 0.29% in the Cell− and Control groups, respectively. Ileal microbiome of Cell+ group was the second highest in abundance of Clostridium sensu stricto compared with Cell− and Control groups, with relative abundances of 31.27%, 0,09%, and 3.37%, respectively. STB and ileal microbiome from Cell+ group were dissimilar to Cell− and Control groups (P < 0.05). Through bacterial isolation, C. septicum and C. perfringens were isolated from SBT samples elucidating a potential synergistic effect in the development of the disease. Additionally, the high abundance of Clostridium spp. in the ileum and SBT provides insight on the potential translocation of Clostridium spp. from the intestine to subcutaneous breast tissue resulting in cellulitis.

Key Words: cellulitis, Clostridium spp., translocation, gut health.

Authors: D. Ayala*, D. Grum, E. Kimminau, N. Evans, K. Russo, B. Trible, and T. P. Karnezos / Purina Animal Nutrition Center, Gray Summit, MO, USA.

M. Trombetta, The Ohio State University

Necrotic enteritis is an enteric disease primarily caused by overgrowth of Clostridium perfringens (CP) in the small intestine following a variety of predisposing factors. The objective was to determine if a novel probiotic showed anti-clostridial effects, survived pelleting temperatures and harsh environment of the gastrointestinal tract (GIT), and if anti-clostridial effects were retained through the GIT. The probiotic was tested against 8 strains of CP to determine overarching anti-clostridial effect. The probiotic suppressed all 8 strains of CP significantly (P <0.05) when CP inoculated media was overlaid onto a pregrown colony of probiotic and zones of inhibition measured. Next, probiotic efficacy was compared against common antibiotics and commercial probiotics. Two antibiotics, penicillin (0.0625 mg/mL) and metronidazole (0.05 mg/mL), both commercial probiotics, and the experimental probiotic reduced CP growth with the experimental probiotic outperforming both commercial probiotics (P < 0.001) and metronidazole (P = 0.007). The CP strain showed resistance to the third antibiotic, BMD (0.022 mg/mL). A germination and sporulation assay was run to ensure the spores could survive pelleting. A lack of significant change (P= 0.112) in cell recovery was indicated the probiotic’s ability to endure pelleting. A simulation digestive assay was performed mimicking the crop, proventriculus, and intestines to ensure the probiotic could survive digestion. When spores recovered from each section of the GIT were compared, the final concentration was significantly lower than the initial (P = 0.018) with a 2-fold reduction reaching the small intestine. The digestive assay was repeated with the addition of CP in the small intestines to determine if anti-clostridial properties were maintained. Using a 10⁶ dose of spores, the reduction in CP was significant at P= 0.103. The results of the experiments indicate the probiotic is a candidate for treatment and control of necrotic enteritis due to its broad anti-clostridial properties and resilience in harsh environments.

Key Words: probiotic, poultry, necrotic enteritis, antibiotic alternatives, direct-fed microbial.

Authors: M. Trombetta *1, K. McGovern 1, A. Duff 1, H. Xue 2, D. Wang 2, L. Johnston 2, and L. Bielke 1 / 1 The Ohio State University, Wooster, OH, USA, 2 Amlan, Chicago, IL, USA.

Chairperson: Mike Kogut / Speaker: Mick Bailey, University of Bristol

There are clear associations between colonization with intestinal microbiomes and the development of the mucosal immune system and of metabolic function. In laboratory rodents, single microbial species can expand whole sections of the immune system and colonization of gnotobiotic pigs with an oligobiota has similar effects. However, in the field, young animals rapidly become colonized with complex microbiomes acquired from their mothers and from the environment. These are a complex set of spatially-linked microbiomes in which oral, gastric, duodenal, jejunal, ileal, cecal and colonic compartments are sequential ecosystems: each compartment acquires microorganisms from the previous compartment and then contributes microorganisms to the next. Once intestinal microbiomes are considered as a set of sequentially-dependent, complex ecosystems, it is apparent that the strategies we use to manipulate them may need to be similarly complex to provide robust, predictable results over multiple units and management systems. Many trials document the administration of single organisms to weaning or growing pigs resulting in measurable change in some parameters and these effects are interpreted as direct effects of the administered organisms (“probiotics”) on host physiology. However, while this interpretation might be acceptable in a gnotobiotic system, it is less so where the probiotic competes with existing, complex ecosystems. In addition, the documented effects frequently vary between trials or disappear once applied in real-world husbandry systems. We propose that most administered ‘probiotics’ and prebiotics, and of many nutritional interventions act indirectly, by modifying the existing, complex spectrum of sequentially[1]dependent microbiomes. As a corollary, we propose that their effectiveness is dependent on the composition of the existing, complex set of microbiomes. Characterization and manipulation of the current, baseline microbiomes in a specific unit may be necessary before administration of nutritional interventions. In addition, microbial consortia may be more effective in modifying these microbiomes than single organisms.

Key Words: microbiome, ecosystem, probiotic, immunology.

Authors: M. Bailey* and C. R. Stokes, University of Bristol, Bristol, United Kingdom.

Shelby Ramirez, BIOMIN America Inc.

Coccidiosis continues to be a major challenge in poultry production, especially in systems no longer using antibiotics. Many producers implement coccidiosis control programs utilizing combinations of coccidiosis vaccines, coccidiostats, and feed additives. Depending on the modes of action of the feed additive, combining or replacing different strategies may be advantageous. Two feed additives were evaluated: (1) a phytogenic (PHYT) as a replacement of a chemical coccidiostat, and (2) a synbiotic (SYN) as an enhancement of a coccidiosis vaccine. Both studies utilized 800 day-old Cobb 500 chicks randomly allocated to 1 of 4 treatment groups (8 replicate pens per treatment with 25 birds per pen). The first study consisted of a non-challenged control (NCC), a challenged control (CC), CC supplemented with zoalene (ZOA; 125 ppm), or CC supplemented with a PHYT (PHYT; 125 g/MT). The second study consisted of NCC, CC, CC administered a coccidiosis vaccine at placement (VAC), or VAC supplemented with SYN (SYN; 500 g/MT). In each study, birds were raised on used litter and on d 19, 20, and 21, 1 × 10⁸ cfu/bird of Clostridium perfringens was administered via the feed. Data from each study were analyzed independently using GLIMMIX procedure of SAS with significance reported at P < 0.05. The CC resulted in a 9-point (P < 0.05) and 6-point (P < 0.05) increase in FCR compared with the NCC for study 1 and study 2, respectively. In study 1, final body weight and FCR were significantly improved (P < 0.05) in the ZOA- and PHYT[1]supplemented groups compared with the CC, but were similar to each other as well as the NCC. In study 2, supplementation with SYN was able to improve FCR by 7 points compared with VAC group (P < 0.05) and was similar to the NCC. Body weight was numerically (P > 0.05) improved in SYN birds compared with VAC birds. Additionally, oocyst shedding was increased (P < 0.05) in VAC and SYN on d 14 and d 21, suggesting normal cycling of the vaccine oocysts had occurred. Incorporating either of these feed additives was advantageous in the context of each coccidiosis control program.

Key Words: coccidiosis, feed additive, broiler.

Authors: S. M. Ramirez *1 , G. R. Murugesan 1, C. M. Pender 1, and B. Lumpkins 2 / 1 BIOMIN America Inc., Overland Park, KS, USA, 2 Southern Poultry Feed and Research Inc., Athens, GA, USA.

Mohammed el Amine Benarbia, NOR FEED

Health in general—gut health in particular—is an important pillar of animal welfare. Gut health can be affected negatively in broiler chickens by coccidiosis. To limit the effect of this disease, coccidiostats have been used with success for decades. However, their intensive use in broiler chicken flocks has led to resistance. Moreover, societal demand for an antibiotic-free animal product is increasing. Thus, there is a need to develop natural and efficient tools to support modern poultry producers to fulfil the productivity needs along with market demand. Saponin[1]rich plants like Yucca schidigera and Trigonella foenum-graecum are promising tools. The objective of this presentation is to share the different evaluation methods applied to assess the efficacy of the saponin-rich plant-based solution Norponin XO 2 (NPXO2) and draw a clear picture of the available alternatives for gut health managers. From 2016 to 2021, 2 types of experimental design were applied. The first one consisted of experimental infestations within research facilities. In this experimental design, 4 groups of birds were used: infested-untreated control (IUC), untreated[1]uninfested control (UUC), infested-ionophore treated (positive control group), and infested-NPXO2 treated (NPXO2). The second experimental design was used in commercial farms with 2 groups, one with “conventional” coccidiosis management tools and one supplemented with NPXO2. In both designs, production performance parameters were monitored and gut health was assess using the Johnson and Reid methodology to score intestinal lesion score (ILS) related to coccidiosis infestation. Results from the first experiment showed that both ionophore and NPXO2 supplementation were able to reduce Eimeria spp. infestation-related ILS and maintain zootechnical performance. In the second experiment, in addition to the fact that NPXO2 supplementation was as efficient as conventional tools, NPXO2 birds displayed a higher livability. These results suggest that NPXO2 supplementation is a reliable tool to be included in gut health management in broiler chicken flocks.

Key Words: gut, Eimeria, chicken.

Authors: M. el Amine Benarbia*, P. Engler, L.-S. Druhet, and P. Chicoteau / Nor Feed, Beaucouzé, France.

Don Ritter, Innovad USA

Coccidiosis in modern production has been linked with compromised intestinal health, chronic inflammation, reduction in antibiotic usage and economic losses. Even subclinical disease can lead to significant production losses. The use of anticoccidials; for example, ionophores and synthetics, has been associated with resistance, whereas vaccines have had various degrees of efficacy. Here we evaluated a novel phytogenic technology (Aflocox, Innovad) with or without an intestinal health enhancer technology (Lumance) (1) under a severe mixed coccidiosis infection in a battery cage setup, and (2) in a dietary-induced mild coccidiosis within a real production system. In experiment 1, a mixed Eimeria infection (E. acervulina ~130,000, E. maxima ~39,000, E. tenella ~32,000 and E. mitis ~11,000 oocysts/mL) at d 13 resulted in 25% mortality between d 19 and 21, which was reduced to 0% by Aflocox (200g/ton from d 13 onward) in combination with Lumance (d 1-d 12: 250 g/T; d 13 onwards: 1 kg/ton) (P < 0.05, n = 4 cages/treatment, n = 5 birds/treatment). In experiment 2, under farming conditions, a high non-starch polysaccharides (NSP) diet (60% wheat + 5% rye without NSPases) increased total Eimeria lesion scores at d 21 (P = 0.09) and significantly worsened macroscopic dysbacteriosis both at d 21 and d 28 over a standard diet (50% corn + 30% soy) (P < 0.005; 1 bird per pen scored, n = 8 pens/group; n = 30 birds/pen), evaluated according to Teirlynck et al. (2011). Both Aflocox and Aflocox in combination with Lumance (a) ameliorated dysbacteriosis over the NSP-challenged group and showed no difference over the standard diet at d 21 and d 28 (P < 0.005), and (b) reduced numerically total Eimeria lesion scores (1 bird per pen scored, n = 8 pens/group; n = 30 birds/pen) and OPGs, to the same level as the standard diet at d 21 and d 28. Importantly, Aflocox (200 g/ton) with or without Lumance (1 kg/ton) at the end of the life cycle (d 35) significantly increased the BW over the high-NSP and reduced the FCR (P < 0.05; n = 8 pens/group; n = 30 birds/pen). Thus, we have evaluated a novel phytogenic technology (Aflocox) against 2 setups of coccidiosis with great promise.

Key Words: coccidiosis, gut health, phytogenic, real farming conditions.

Authors: A. Khadem 1,2, D. Ritter *3, and C. Gougoulias1 / 1 Innovad NV/SA, Essen, Belgium, 2 Lab of Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium, 3 Innovad USA, Salisbury, MD, USA.

Glenn Zhang, Oklahoma State University

Necrotic enteritis (NE) is known to induce intestinal dysbiosis. However, the intestinal microbes that are associated with NE severity are yet to be characterized. Here, we investigated the link between the ileal microbiota/mycobiota and disease severity in a chicken model of clinical NE using 16S rRNA gene and ITS2 sequencing. Our results indicated that, while the total bacterial population in the ileum was increased by 2- to 3-fold in NE chickens, the total fungal population progressively declined in exacerbated NE, with the most severely infected chickens showing a nearly 50-fold reduction relative to mock-infected controls. The C. perfringens population was increased from 0.02% in healthy chickens to 58 to 70% in chickens with severe infection, with most the ileal microbes being markedly diminished. Compositionally, a large group of ileal microbes showed a significant positive or negative correlation with NE severity. Firmicutes, such as group A and B Lactobacillus, Lactobacillus reuteri, Subdoligranulum variabile, Mediterraneibacter, and Staphylococcus as well as 2 genera of Actinobacteria (Corynebacterium and Kocuria) and 2 closely related Cyanobacteria were progressively declined as NE was aggravated. Other Firmicutes, such as Weissella, Romboutsia, Kurthia, Cuneatibacter, Blautia, and Aerococcus, appeared much more sensitive and were rapidly abolished in chickens even with mild NE. On the other hand, Enterococcus cecorum and 2 Escherichia/Shigella species were only enriched in the ileal microbiota of chickens with extremely severe NE. Multiple fungi such as members of the Wallemia and Aspergillus species were obviously diminished in response to NE. Functionally, secondary bile acid biosynthesis was predicted to be suppressed by NE, while biosynthesis of aromatic and branched-amino acids and metabolism of most amino acids were enhanced. These intestinal microbes showing a strong correlation with NE severity may provide important leads for the development of novel diagnostic or therapeutic approaches to NE and possibly other enteric diseases.

Key Words: necrotic enteritis, microbiome, mycobiome, dysbiosis, Clostridium perfringens.

Authors: Q. Yang and G. Zhang*, Oklahoma State University, Stillwater, OK, USA.

Hongyu Xue, Amlan International

M52 is a natural anticoccidial feed additive that has been shown to help prevent coccidiosis, maintain bird productivity, and has potential as an alternative to ionophores and chemical coccidiostats. This study further evaluated the effects of the product on host anti-Eimeria immunity, gut microbiome composition and intestinal integrity of broilers challenged with experimental coccidiosis. In this 28-d study, day-old Cobb broiler chicks were randomly assigned to 1 of 3 groups: (1) unchallenged control; (2) Eimeria-infected control; and (3) Eimeria-infected + M52 (70 g/MT feed). On d 14. chickens from treatments 2 and 3 were challenged with 100X Bio-Coccivet R Vaccine, a live polyvalent vaccine consisting of sporulated oocysts of 7 Eimeria spp. Peripheral blood mononuclear cell phenotype, ceca-cecal tonsil cytokine mRNA expression, duodenal/jejunum histopathology and gut microbiome of cecal content were examined. Eimeria challenge induced moderate to severe parasitic enteritis manifested with prominent villous hyperplasia, heterophil mucosal infiltration, and hemorrhagic or necrotic foci on d 19. The product markedly reduced these histopathological changes. On d 28, M52 significantly increased the abundance of CD4−TCRVβ1+, CD8−CD28+ and CD4−TCRVβ1+ subsets, which help uphold mucosal immune homeostasis and develop competent mucosal and systemic adaptive immunity when faced with pathogen insults. On d 19, M52 dampened Eimeria challenge-associated upregulation of cecal IL-10, whose immunosuppressive properties can also be exploited by pathogens to facilitate their own survival. Before Eimeria challenge, M52 enhanced the relative abundance of Blautia and L-Ruminococcus genera, 2 short-chain fatty acid producers, and further dampened the impairment on microbiota diversity associated with the subsequent Eimeria challenge. Collectively, M52 treatment promoted a well-balanced immune homeostasis, dampened intestinal damage, and preserved the microbiota diversity, which all contribute to an enhanced resilience to Eimeria spp. challenge.

Key Words: coccidiosis, anti-Eimeria immunity, IL-10, micro[1]biome, intestinal integrity.

Authors: H. Xue *1, F. Rigo 2, B. Beirão 2, C. Fávaro 2, and M. Ingberman 2 / 1 Amlan International, Chicago, IL, USA, 2 Imunova Análises Biológicas, Curitiba, Paraná, Brazil.

A. J. Calderon, Auburn University

Young piglets are more susceptible to severe immune challenges because their immune system is not fully developed at birth. A randomized complete block design experiment utilizing a 4 × 2 factorial treatment arrangement was conducted to evaluate the effect of dietary protein source on intestinal crypt cell proliferation and local macrophage populations of neonatal piglets. Male and female piglets (n = 123) from 5 farrowing groups were fed 1 of 3 liquid diets: (1) Whey (MLK) only, (2) MLK + spray-dried plasma (SDP), or (3) MLK + soy protein concentrate (SPC). After receiving sow colostrum, piglets were reared in Rescue Decks® until tissue collection. A fourth group was left in the farrowing crate and nursed the sow until sampling (SOW). Duodenal (DUO), jejunal (JEJ), and ileal (ILE) tissue was collected at 6, 14, 19, and 25 d of age. One hour before euthanasia, pigs were injected with 5′-bromo-2′-deoxyuridine (BrdU) to label proliferating (BrdU+) cells. Proportion of proliferating (BrdU+), crypt epithelial cells (PCEC) and CD172a (total) and CD80+ (pro-inflammatory) macrophage populations was assessed after cryohistology and immunofluorescence staining. At 6 d old, male piglets fed SDP had greater JEJ PCEC compared with SOW males (P = 0.0081). At d 14, male piglets fed SPC had higher DUO PCEC than those fed SDP (P = 0.0074). Females fed SPC tended to have greater JEJ (P = 0.0894) and ILE (P = 0.0793) PCEC than SPC-fed males. At d 6, male SOW-fed piglets had the largest populations of both DUO pro-inflammatory (CD80+; P= 0.0478) and proliferating, pro-inflammatory (BrdU+:CD80+; P = 0.0040) macrophages. In the DUO, at d 19, SDP-fed and male SPC-fed piglets exhibited the highest total macrophage density (P = 0.0025). In the ILE, on d 19, SDP-fed females had the highest pro-inflammatory macrophage density compared with all other groups (P = 0.0465). These data establish a model to evaluate developmental intestinal innate immunity and indicate that dietary protein source can impact PCEC, which may benefit renewal of the intestinal epithelial barrier.

Key Words: immune system, small intestine, intestinal macro[1]phages, spray-dried plasma.

Authors: A. J. Calderon*, J. L. Sandoval, J. D. Starkey, and C. W. Starkey, Auburn University, Auburn, AL, USA.

Famata Perry, University of Delaware

The aim of this study was to evaluate the effectiveness and mechanism of action of 2 feed additives in reducing the impacts of physiological stressors. We compared the effects of protected biofactors and antioxidants [P(BF+AOx)], and protected biofactors and antioxidants with protected organic acids and essential oils [P(BF+AOx)+P(OA+EO)] on the immune and metabolic health of Ross 308 chickens. These biofactors and antioxidants were derived from vitamins, and Aspergillus niger, Aspergillus oryzae, and Bacillus subtilis fermentation extracts. All Ross 308 chickens were exposed to a double dose of live bronchitis vaccine at d 0 and environmentally challenged by reducing the temperature from 30–32°C to 20–23°C at d 3 for 48 h. Control birds were fed without feed additives in the diet. Performance data, jejunum and liver samples were collected to evaluate the effects of these treatments on growth, cytokine expression and kinome peptide array. Analysis of variance was used for statistical analysis of the performance and gene expression data (P = 0.05), and PIIKA2 was used for statistical evaluation and comparison of the kinome peptide array data. The P(BF+AOx) and P(BF+AOx)+P(OA+EO) treatments significantly increased bird weight gain and feed conversion ratio. The kinome peptide array data showed increased activity of cytoskeletal, cell growth and proliferation proteins, and metabolic signaling in the jejunum of P(BF+AOx)+P(OA+EO) treated chickens. There was also increased phosphorylation and activity of proinflammatory and immune proteins in the liver compared with the jejunum. There was a significant decrease in IL-6 gene expression in the jejunum of P(BF+AOx)+P(OA+EO) samples compared with control at d 15, and a decreased expression of the anti-inflammatory marker IL-10 for P(BF+AOx)+P(OA+EO) and P(BF+AOx) liver groups when compared with control at d 7. P(BF+AOx)+P(OA+EO) improves gut health via growth and metabolic signaling in the jejunum while both treatments induce immunomodulation.

Key Words: biofactor, antioxidant, immunometabolism, kinome peptide array, gut health.

Authors:  F. Perry *1, L. Lahaye 2, E. Santin 2, C. Johnson 1, D. Korver 3, M. Kogut 4, and R. Arsenault 1 / 1 University of Delaware, Newark, DE, USA, 2 Jefo Nutrition Inc., Saint-Hyacinthe, QC, Canada, 3 University of Alberta, Edmonton, AB, Canada, 4 USDA-ARS, Southern Plains Agricultural Research Center, College Station, TX, USA.

Cristiano Bortoluzzi, DSM Nutritional Products AG

The objective of the present study was to evaluate the complete metagenomics and metabolomics of the cecal microbiome as well as the intestinal transcriptomics of broiler chickens supplemented with a novel precision glycan Microbiome Metabolic Modulator (MMM). Day-old chicks were placed on a completely randomized block design with 2 treatments, 21 replicates, and 40 birds/replicate. The treatments consisted of a non-supplemented control or feed supplemented with 500 ppm of MMM2 precision glycan (Glycan M2–1, Midori USA, Inc., Cambridge, MA, USA; commercially available as Symphiome Poultry; DSM Nutritional Products). Cecal content samples were collected at 24 and 42 d of age from 1 bird/pen, and frozen at −80°C for further analyses. Ileal tissue samples were collected at 42 d of age into RNAlater solution and frozen at −20°C until RNA extraction. Metagenomic DNA from the cecal content was isolated and sequenced by an Illumina HiSeq 3000 platform, and the entire metabolomics analysis was performed. Ileal mRNA was isolated, and the gene expression analysis was performed by a chicken Gene expression Microarray, 4x44K (Agilent). Related to the microbiome analysis, increased energy production around the TCA cycle associated with a concomitant increase of acetyl[1]CoA, justified mainly by a significant acetylation of multiple amino acids and increased lipid biosynthesis was observed. The increased carbon flow was also directed to the production of short chain fatty acids, mainly propionate by the acrylate pathway. About the nitrogen metabolism, ammonia was found to be largely detoxified through pathways to polyamines, out of the uric acid cycle at d 24, and through the asparagine outlet at 42 d. Interestingly, host glutamine synthetase was significantly decreased suggesting decreased ammonia toxicity. Regarding the host meta-transcriptomics, it was shown that IL-4 and IL-4-like, as well as IL-10 family were positively associated with MMM2 supplementation, indicating a possible anti-inflammatory and/or regulatory role of MMM2, most likely as a secondary outcome from the metabolic modulation of the microbiome.

Key Words: broiler, microbiome metabolic modulator, nitrogen metabolism.

Authors:  J. Claypool 1, K. Freeman 1, B. Blokker 2, M. C. Walsh 3, C. Bortoluzzi *3, and G. Schyns 1,3 / 1 DSM Nutritional Products Ltd., Lexington, MA, USA, 2 CRNA, DSM Nutritional Products SA, Village-Neuf, France, 3 DSM Nutritional Products AG, Kaiseraugst, Swittzerland.

Stephanie Hutsko, Provimi

Coccidiosis in turkeys leads to decreased ADG, increased FCR, damaged intestinal integrity and, in clinical cases, increased mortality. Over the past decades of chemical and medication usage, Eimeria species have developed resistance to anticoccidial drugs limiting their effectiveness. So, this creates an urgent need to develop effective prevention and control strategies. Two studies were conducted to evaluate the potential of a proprietary saponin/ oregano blend to maintain performance and reduce oocyst shedding in turkey poults during a coccidiosis challenge. In study 1, poults were raised in battery cages d 0- 14 and then were randomly assigned to 1 of 4 treatments: Control (C; non-medicated, non-infected), Negative Control (NC; non-medicated, infected), Zoalene (ZO; 125 ppm Zoalene, infected) and Experimental (E; saponin/oregano blend; 2 lb/t, infected), each with 7 replicates of 8 poults/replicate. On d 16 poults were gavaged with 1 mL of coccidia inoculum (100,000 oocysts/mL) and on d 25 birds were weighed and excreta collected for oocyst counts. The E birds had higher ADG compared with the NC and ZO birds and a lower FCR compared with NC (P < 0.05). Shedding of E. adenoids was decreased in the E group compared with NC, and shedding of E. meleagrimitis and E. gallopavonis was intermediate between the NC and ZO groups. In study 2, poults were again raised in battery cages d 0–14 and then were assigned to 1 of 5 treatments: Control (C; non-medicated, non-infected), Negative Control (NC; non-medicated, infected), Zoalene (ZO; 125 ppm, infected), Natustat (NS; 4lb/t, infected), Experimental (E; saponin/oregano blend; 2 lb/t, infected), each with 7 replicates of 8 poults/replicate. On d 16 poults were gavaged with 1 mL of coccidia inoculum (100,000 oocysts/mL). On d 21, birds were weighed and excreta collected for oocyst counts. Birds in the E group showed improved ADG and FCR compared with the NC and NS groups (P < 0.05). Oocyst shedding in the E group was intermediate between the NC and ZO groups. Overall, this saponin and oregano blend was effective at improving performance and reducing oocyst shedding in poults during a coccidiosis challenge.

Key Words: cocci, prevention, turkey, oocyst, saponin.

Authors: S. Hutsko*, A. Garcia, E. Guaiume, D. Giesting, and R. Payne / Cargill Animal Nutrition, Rice Lake, WI, USA.

Abdullah Mahfuz, Feed Energy Company

Due to the recent restrictions of antibiotic, antibiotic as growth promoter (AGP) usage in animals and the negative effects of formaldehyde on animal performance, the animal feed industry has started to evaluate alternatives to AGP and formaldehyde for flock health and performance. R2 is Feed Energy’s patent pending low pKa, lipid-based line of products that provides nutrient-dense source of essential fatty acids along with feed biosecurity benefits. The objective of this study was to determine the effects of R2 product OptimizR2 on broiler health and performance in comparison with commonly used antimicrobial measures. One-day-old male Ross 308 chicks (n = 320) were placed in 40 cages of 8 birds/cage. The treatments consisted of feeds formulated to meet NRC with 4% added lipids to produce 4 dietary treatments (SBO: Basal Diet+Refined SoyOil, SBO-B: SBO+Bacillus product, SBO-F: SBO+Formaldehyde product, Basal Diet+OptimizR2). Treatments were randomly assigned to provide 10 replications/treatment. The birds were fed a starter and grower feeds from d 1 to d 21 and d 22 to d 35, respectively. Microbial phylogenetic diversity analysis of d-35 cecal samples of birds were conducted using 16S rRNA gene sequences. Performance parameters measured were body weight (BW), average daily gain (ADG), feed conversion (FC), and mortality. Differences in performance between groups were analyzed for significance (P < 0.05) using ANOVA. The numerical differences in BW (SBO = 4.30 lb, OptimizR2 = 4.50 lb, SBO-B = 4.33 lb, SBO-F = 4.19 lb) and ADG (SBO = 0.12, OptimizR2 = 0.13, SBO-B = 0.12, SBO-F = 0.12) at the end of the trial were different (P < 0.05). The dietary addition of R2 to the broiler diets also improved FC (P < 0.05). Cecal microbiota showed much higher diversity with beneficial bacteria in R2 group. R2 preserve higher population of lactobacillus and eliminate harmful gram-negative bacteria compare with bacillus group. Gut beneficial lactobacillus level was low in formaldehyde group. The addition of R2 to feeds improves performance and offers benefits to the microbiome in broilers.

Key Words: broiler, microbiome, nutrition.

Authors: A. Mahfuz *1, C. Jaeger 1, E. Weaver 2, and M. Dasari 1 / 1 Feed Energy Company, Pleasant Hill, IA, USA, 2 South Dakota State University, Brookings, SD, USA.

Haitham Yakout, Adisseo

Aflatoxins are hepatotoxic and carcinogenic, and display immunosuppression properties for both humans and animals. This is also why they are the most widely studied and regulated mycotoxins. Strategies to mitigate aflatoxins prevalence includes the use of clays in animal feed as sequestering agents. Innovative approach consists in providing ingredients controlling inflammation and promoting immune. The objective of this study is to evaluate the impact of naturally contaminated diet with aflatoxins (AFB1) on health and performance of commercial broilers, and the efficacy of a mycotoxin deactivator based on clays, inactivated yeast and fermentation extracts. A total of 7200 straight run Vencobb 430 chicks were assigned for 42 d to 1 of 3 treatments in a randomized block design (8 pens per treatment): (1) BD: basal diet with residual mycotoxins; (2) MT: diet with 56 ppb AFB1; and (3) TN: MT + mycotoxin deactivator at 2 kg/t. Performance parameters were recorded on weekly basis, and blood was collected at d21 and d42. MT group shows significant lower BW than BD from 14 to 42 d, resulting in a decreased FCR (+2.6% for the overall period). TN can restore the performance compared with MT (−9.7% FCR) and to BD (−7.1% FCR). AFM1 in plasma, as a biomarker of chronic exposure, showed reduction of AFB1 assimilation (from −42% to −75%, P < 0.01) in the TN treated birds. AFB1 induces inflammation as shown by the increased secretion of pro-inflammatory cytokines such as IL1b, IL6, IL8, TNFa between BD and MT. TN treatment aimed to decrease these parameters relative to MT and to promote the production of anti-inflammatory IL10. Birds receiving TN treatment have higher antioxidant indicators such as superoxide dismutase, glutathione and glutathione peroxidase compared with MT and showed lower lipid peroxidation marker like malondialdehyde. This study highlights the complementary modes of action of this mycotoxin deactivator by inhibiting AFB1 in the intestine and by repairing damage caused at the gut level on inflammation, immune response and redox balance. This leads to sustained broiler resilience and performance.

Key Words: aflatoxin, broiler, immunity, inflammation, antioxidant.

Authors: P. S. Ingewar 1, V. Patil 2, N. Kurkure 2, G. Bromfman *3 , H. Yakout 3, J. Dvorska 4, and D. P. Preveraud 4 / 1 A2 Livestock Farms and Research, Nagpur, India, 2 Department of Pathology, Nagpur Veterinary College, Maharashtra Animal & Fishery Science University, Nagpur, India, 3 Adisseo USA Inc., Alpharetta, GA, USA, 4 Adisseo France SAS, Antony, France.

Chairperson: Cristiano Bortoluzzi / Speaker: Maria Mendoza, Evonik Corporation

The functional amino acids (FAA) Met, Thr, and Trp play an important role supporting the immune system and gut health. Two studies were designed to evaluate the supplementation of FAA above requirements during a Salmonella Typhimurium (ST) challenge. Experiment 1 was conducted for 14 d in a 2 × 2 × 2 factorial arrangement, consisting of 2 CP levels (16% or 20%), 2 FAA levels (Met, Thr, and Trp at 100% [FAA−] or 120% of requirements [FAA+]), and 2 challenge conditions (saline [CT] or ST inoculation). Pigs (initial BW = 13.9 kg; 8 pigs/treatment [Trt]) received the diets from d 0 to 14, and on d 7 pigs were inoculated. Pigs inoculated with ST had higher rectal temperature, serum haptoglobin, and activity of antioxidant systems, softer feces, and lower growth compared with CT pigs. Pigs challenged with ST and fed FAA+ diets had a less severe acute inflammatory response and greater growth compared with FAA− counterparts. Pigs fed 16% CP diets had lower cecal ST score compared with pigs fed 20% CP diets. Protein level did not affect growth and immune status. Experiment 2 was conducted for 21 d to evaluate whether providing an adaptation period of FAA+ is beneficial during health challenges. All pigs (initial BW = 11 kg, 8 pigs/trt) were inoculated with ST on d 14. Pigs were fed FAA− or FAA+ diets at different periods. The 4 treatments included (1) FAA−, only FAA−, (2) FAA+15, pigs received FAA− and from d 15 FAA+, (3) FAA+8, pigs received FAA− and from d 8 FAA+, and (4) FAA+, only FAA+. As in Exp. 1, pigs experienced an acute inflammatory response due to ST regardless of the treatments. Pigs fed FAA+ diets had lower levels of serum haptoglobin, activity of antioxidant systems, cecal ST score, and greater growth compared with FAA−. Pigs fed FAA+15 and FAA+8 had intermediate activity of antioxidant systems and growth compared with FAA− and FAA+. Both experiments demonstrated that pigs fed FAA+ are better equipped to counteract an ST infection while maintaining optimum growth, and the benefits are greater when pigs are fed FAA+ diets for a longer period.

Key Words: functional amino acids, pig, Salmonella.

Authors: L. A. Rodrigues 1,2, M. O. Wellington 2, J. C. González-Vega 3, J. K. Htoo 3, A. Menconi 4, S. M. Mendoza *4 , A. G. Van Kessel 2, and D. A. Columbus 1,2 / 1 Prairie Swine Centre Inc., Saskatoon, Saskatchewan, Canada, 2 Department of Animal and Poultry Science, University of Saskatchewan, Saskatoon, Saskatchewan, Canada, 3 Evonik Operations GmbH, Hanau-Wolfgang, German, 4 Evonik Corporation, Kennesaw, GA, USA.

Yang Lin, University of Georgia

A total of 252 Cobb 500 male broiler chicks were used in a 21-d experiment to study the possibility of xylo-oligosaccharides (XOS) helping to recover gut impairment in Eimeria-challenged broilers by regulating the expression of nutrient transporters and tight junctions, and cecal short-chain fatty acids (SCFA), which is an indicator of bacterial status. Birds were allocated to 6 treatments in a 3 × 2 factorial arrangement (3 corn-soybean diets with 0, 0.5, 1 g/kg XOS × with or without Eimeria challenge). Each treatment had 6 replicates with 7 birds per replicate. Challenged groups were inoculated with a solution containing E. maxima, E. tenella, and E. acervulina oocysts on d 15. On d 21, jejunal tissue was collected for gene expression analysis and cecal content was collected for SCFA analysis. The Eimeria × XOS interaction for tight junction claudin 1 showed that both 0.5 and 1 g/kg XOS alleviated (P < 0.05) Eimeria-induced claudin 1 upregulation. The Eimeria × XOS interaction for sugar transporters showed the extent of Eimeria-induced GLUT2 and GLUT5 downregulation was smallest in the 0.5 g/kg XOS supplemental treatment. In addition, Eimeria upregulated (P < 0.01) tight junction JAM2 and glucose transporter GLUT1 but downregulated (P < 0.01) the peptide transporter PepT1, amino acid transporters rBAT, CAT2, y+LAT2, and zinc transporter ZnT1. Eimeria decreased (P < 0.05) cecal saccharolytic SCFA acetate, butyrate and total SCFA, but increased (P < 0.05) cecal branched-chain fatty acids isobutyrate and isovalerate. The supplementation of XOS tended to decrease the concentration of isobutyrate (P = 0.080) and isovalerate (P= 0.062). In conclusion, Eimeria challenge triggered changes in expression of tight junction and nutrient transporter genes. Supplemental XOS helped reverse the gene expression changes in tight junction claudin 1 and glucose transporter GLUT2 and GLUT5, and showed the potential of alleviating the Eimeria[1]induced unfavorable cecal fermentation pattern.

Key Words: xylo-oligosaccharides, Eimeria, tight junction, nutrient transporter, short-chain fatty acids (SCFA).

Authors: Y. Lin* and O. Olukosi, University of Georgia, Athens, GA, USA.

Gilson Gomes, AB Vista

Salmonella control in poultry flocks and its public health impact is crucial. The objective of this study was to evaluate the effects of a stimbiotic (Signis, AB Vista, Marlborough, UK). A total of 900 day-old chicks were randomly allocated to 60 pens. The trial design was a complete 2 × 2 × 2 factorial with 2 based diets, a standard commercial diet with (PC) or without a multi NSP enzyme and a prebiotic (NC), with or without the stimbiotic at 100 g/ton of feed fed to males or females. Body weight (BW), feed intake (FI), and livability were recorded per pen. The feed conversion ratio was corrected for mortality (mFCR) and BW (mbwcFCR). Four birds per pen were randomly selected and cecal content collected and pooled for the enumeration of Salmonella by most probable number (MPN) test culture. Performance data were submitted to a 2-way ANOVA, means were separated using a Student’s test with significance accepted when P ≤ 0.05. Livability and Salmonella MPN were analyzed by Chi-squared where means were separated using Wilcoxon test. Contingency analysis was performed on the presence/absence % of Salmonella. No effect was observed on livability. For almost all criteria tested there was an interaction between the stimbiotic and sex, which means in this study a greater effect in males. An interaction between diet and stimbiotic was noticed on Salmonella count (P < 0.05), which was reduced for birds fed the stimbiotic, but this effect was more pronounced when supplemented on top of the NC. As a result, the proportion of Salmonella positivity was reduced from 26.7% to 13.3% when the stimbiotic was supplemented regardless of the diet. There was an interaction between the diet and the stimbiotic on mFCR (P < 0.05), mbwcFCR (P < 0.05), and FI (P < 0.05), with a larger effect noticed when supplemented to NC. In conclusion, birds fed the stimbiotic showed better performance while reducing Salmonella positivity, irrespective of sex and diet. Thus, the stimbiotic can serve as a nutritional strategy to improve performance and gut resilience mitigating Salmonella proliferation in the intestinal tract.

Key Words: stimbiotic, Salmonella, gut resilience, performance.

Authors: X. Rousseau*, N. Amornthewaphat, G. A. Gomes, and T. T. dos Santos, AB Vista, Marlborough, United Kingdom.

S. F. Leiva, Auburn University

Previous studies demonstrate that maternal supplementation of the circulating metabolite of vitamin D3 (D3), 25-hydroxycholecalciferol (25OHD3), enhances the immunocompetence of broiler chick offspring. To assess the effect of combining maternal (MD) and post-hatch (PD) dietary 25OHD3 inclusion on duodenal (DUO) crypt cell proliferation and local innate immunity of young broiler chickens, a randomized complete block design experiment with a 2 × 2 factorial treatment structure was conducted. All diets were formulated to provide 5,000 IU of vitamin D. From 25 to 41 wk-of-age, broiler breeder hens were fed 1 of 2 MD: 5,000 IU D3 per kg of feed (MCTL) or 2,240 IU of D3 + 2,760 IU of 25OHD3 per kg of feed (M25OHD3). Male broiler offspring (n = 480) hatched from eggs collected from 41-wk-old hens were fed 1 of 2 starter PD: 5,000 IU D3 per kg of feed (PCTL) or 2,240 IU D3 + 2,760 IU 25OHD3 (P25OHD3). DUO samples (n = 12 birds per treatment per day) were collected on d 3, 6, 9, 12, 15, 18, and 21 for cryohistological and immunofluorescence analysis to facilitate the enumeration of total macrophages, CD80+ macrophages (pro-inflammatory macrophages), and mitotically active cells (BrdU+) to calculate the proportion of proliferating cells per duodenal crypt (PPC). Broilers from the M25OHD3:PCTL group had higher PPC than MCTL:PCTL on d 3 (P = 0.002). On d 9, MCTL:P25OHD3 broilers had more proliferating, proinflammatory macrophages compared with those from the M25OHD3:P25OHD3 treatment (32 vs. 17 cells per mm2; P = 0.023). M25OHD3:PCTL broilers had more total proliferating macrophages than M25OHD3:P25OHD3 birds (89 vs. 56 cells per mm2; P = 0.033). At d 21, a MD × PD interaction was observed, where birds from the M25OHD3:P25OHD3 treatment had more proliferating macrophages than M25OHD3:PCTL broilers (P = 0.029). In conclusion, these results indicate that combined MD and PD 25OHD3 supplementation may alter early post-hatch duodenal development and local intestinal innate immunity parameters.

Key Words: 25-hydroxycholecalciferol, vitamin D, intestinal de[1]velopment, intestinal macrophages.

Authors: S. F. Leiva *1, L. P. Avila 1, G. A. Abascal-Ponciano 1, J. J. Flees 1, K. M. Sweeney 2, J. L. Wilson 2, J. D. Starkey 1, and C. W. Starkey 1 / 1 Department of Poultry Science, Auburn University, Auburn, AL, USA, 2 Department of Poultry Science, University of Georgia, Athens, GA, USA.

K. D. Teague, University of Arkansas

The objective of this experiment was to investigate the effects of the soy galactooligosaccharides (GOS) raffinose and stachyose on performance and gastrointestinal health in young broilers. Birds were fed a GOS-devoid diet based on soy protein isolate (SPI) or the SPI diet with 0.9, 1.8, 2.7, or 3.6% GOS. Stachyose and raffinose were maintained at a ratio of 4:1, similar to what is found in soybean meal, and were added at the expense of corn starch. These 5 treatments were administered to 10 replicate battery cages of 8 birds (0.04 m²/bird). Birds had ad libitum access to feed and water and BW and feed consumption were recorded on 0, 7, 14, and 21 d post-hatch for calculation of feed intake (FI), body weight gain (BWG), and feed conversion ratio (FCR). On 14 and 21 d, whole blood was collected for complete blood counts. Before blood collection on 21 d, birds were orally gavaged with fluorescein isothiocyanate-dextran (FITC-d), and serum samples were analyzed for FITC-d as a marker of gut leakage. Excreta were collected on d 14 for moisture analysis. Crop presumptive lactic acid bacteria (LAB) counts and crop and ceca pH were recorded at d 21. From 0 to 21 d, FI increased linearly (P < 0.01) as dietary GOS increased, whereas BWG increased (P < 0.05) quadratically. Feed conversion ratio increased (P < 0.01) linearly, with a 3-point increase from birds fed SPI to 3.6% GOS. On d 14, there was a linear increase (P < 0.01) in excreta moisture as dietary GOS increased. There was a quadratic increase (P < 0.05) in crop LAB recovery and a tendency for lower (P = 0.08) crop pH as GOS increased, whereas ceca pH decreased (P < 0.01) linearly. At both d 14 and 21, linear increases (P < 0.05) in whole blood heterophil to lymphocyte ratios were observed as dietary GOS increased. Serum concentrations of FITC-d increased quadratically (P < 0.01) as dietary GOS increased, with the greatest numerical FITC-d observed at 1.8%. No histopathological lesions were observed in duodenum or ileum of all groups. Results from this trial indicate that soy GOS have dose-dependent effects on broiler health and feed efficiency from 0 to 21 d.

Key Words: soybean meal, oligosaccharide, raffinose, stachyose.

Authors: K. D. Teague *1, G. Tellez-Isaias 1, V. Petrone-Garcia 2, C.N. Vuong 1, A. Blanch 3, S. H. Rasmussen 3, K. Brown 3, and S. J. Rochell 1 / 1 University of Arkansas, Fayetteville, AR, USA, 2 National Autonomous University of Mexico, Mexico City, Mexico, 3 Hamlet Protein, Horsens, Denmark.