The presence of Salmonella enterica throughout the production and processing continuum is a concern in broiler industry. While federal regulations have lowered the acceptable level of Salmonella contamination on broiler carcasses, the mechanisms that contribute to pathogen attachment are not fully understood. Salmonella Kentucky has become the predominant Salmonella serovar isolated from broilers carcasses at the end of the immersion chill tank. In Europe and Africa this serovar has been shown to acquire antibiotic resistance genes that may lead this non-typhoidal serotype to become a potential public health concern. To investigate the genes that are involved in colonization of the bacteria to broiler skin, a mutant library of the bioluminescent strain of S. Kentucky was constructed. According to the chicken attachment assay, it was concluded that attachment is a multifactorial process with the following elements contributing: i), flagella, ii), LPS structure, iii), amino acid metabolism, iv), TCA cycle pathway; v), conjugative transfer system, vi), multidrug resistant protein, vii), signaling and transportation system, viii), metabolism, ix), different enzymes, x), phage tail fiber protein H, xi), fimbrial export usher proteins, xii), membrane proteins xiii), and several unnamed proteins. The role of flagella between all of these contributing elements appeared to be the most significant. The flagella motor gene, filament sub-units and hook associated protein were deleted by using the ë red recombination method. The mutants’ ability to colonize broiler skin was compared to their parental strain, and the motility and flagellin main sub-unit (FliC) were recognized as the key factors contributing to bacterial attachment. Using Caco-2 cell lines as a cell model to assess adhesion and invasion capacity of flagella mutants, similar results were observed. Based on the result of the experiments conducted in this study, it appears that the active flagella FiC sub-unit plays an important role in colonization of epithelial cells outside and inside of the broilers.

Tunnel ventilation is the design practice of placing exhaust fans and air inlets on opposite ends of animal production facilities and moving air through the building via negative pressure. Increasing air velocity within tunnel ventilated broiler production facilities increases sensible cooling and reduces the need for latent cooling (panting), which improves production efficiency. An air velocity measurement system was developed and measurement density analysis for quantifying air velocity distribution was performed in a 12.19 x 121.9 m commercial broiler production facility. Results showed that axial measurement distances of 3.05 m and 40 measurement points per cross-section produced the most descriptive air velocity distribution maps. Air velocity distribution, mean cross-sectional air velocity, and total facility air flow was assessed in three tunnel ventilated commercial broiler production facilities. These facilities differed in size, design, and equipment configurations (test facility 1 was 18.3 × 170.7 m, test facility 2 was 15.24 × 144.8 m, and test facility 3 was 12.19 × 121.9 m). Air velocity distribution varied within all three facilities. Normalized cross-sectional air velocity was plotted against proportion of total house length to compare the cross-sectional air velocity of the three facilities. House width and physical arrangement of the feed hoppers, heating systems, and tunnel fans are impacting air velocity uniformity and mean cross-sectional air velocity in the three broiler houses. Precision Livestock Farming (PLF) seeks to improve production efficiency and animal well-being by model based control of animal production facilities. Four broiler production facilities were assessed for spatial bird body weight (BW) variability. The facilities were 15.24 × 144.8 m solid side-wall tunnel ventilated broiler houses containing birds at 58 and 59 d of age. Significant differences in BW were found between birds residing at center house (3.47 kg average) and at the side-walls (3.38 kg average) (P = 0.025). This variability in BW could be attributed to any number of environmental, nutrient, or behavioral causes. A discussion of input/control parameters for PLF management of broiler production is presented. Quantification of performance variability within these facilities and defining models for control of input parameters is essential to making PLF management feasible.

Salmonella and Campylobacter are two foodborne pathogens that continue to persist in broiler processing. Various studies have demonstrated that peracetic acid can effectively reduce the prevalence pathogens on broiler meat. However, there are a limited number of studies comparing the effects of peracetic acid on broiler meat from different processing plants. The objective of this study was to determine the prevalence of Salmonella, Campylobacter, and spoilage microbes on broiler meat at different stages of poultry processing in commercial plants that use peracetic acid as the primary antimicrobial. Results indicated that there was a high initial microbial presence on broiler meat at initial stages of processing in all three plants. Peracetic acid effectively reduced the prevalence and microbial load of all microbes analyzed in this study. All microbes were reduced to nondetectable levels in the finishing chiller. However, contamination of all microbes in mechanically deboned meat closely resembled initial carcass contamination. In conclusion, the intervention with the greatest effect on microbial prevalence was peracetic acid in carcass chilling tanks, and, given the level of contamination in mechanically deboned meat, an intervention at this step would be worth investigating.

Currently, feed enzyme supplementation into commercial broiler diets is a routine practice due to hydrolysis of antinutrients, such as non-starch polysaccharides (NSP) and phytate (IP6); ultimately leading to optimized broiler production. As a result, the market has grown rapidly, and a multitude of feed enzymesare commercially available. Complete removal of antinutrients has not been achieved and research has primarily focused on the use of singular enzyme inclusion. Due to biochemical differences among enzyme characteristics, there may be potential for synergistic action to remove antinutrients and further maximize broiler performance. Therefore, the overall objective of this dissertation was to investigate enzyme strategies to determine optimal inclusion for broilers. Chapter 2 investigated the interactive effects of phytase dose (PD; 250 to 1500 FTU/kg) and carbohydrase enzyme (CE) of xylanase (XAN), β-mannanase (MAN), or their combination (XM). These data demonstrated an interaction between CE and PD for d 0-14 FCR, but this significance was lost during the remainder of the study. Overall benefits for performance and processing were demonstrated with feeding a PD of 1500 FTU/kg and XAN. Chapter 3 took a preliminary approach (0-14 d) to investigate the potential synergy among 3 phytase enzymes of varying biochemical properties when fed alone or in combination with low PD (120 or 240 FTU/kg). These data demonstrated potential synergy with the supplementation of 2 phytases combined at a higher PD (240 FTU/kg), as identified from ileal IP (inositol phosphate) lower ester concentration and tibia ash. Chapter 4 built upon the previous chapter, choosing 2 phytases (A or B) and addressing previous limitations by including a broader/more practical range of phytase activity (250 or 1500 FTU/kg); providing 3 diets varying in calcium (Ca) and non-phytate phosphorus (nPP); and using an entire grow-out. Feeding diets lowest in Ca and nPP along with 1500 FTU/kg of A, B, or A+B resulted in improved broiler performance. Additionally, the use of A+B at 250 FTU/kg within diets of medium Ca and nPP levels demonstrated synergism through improved broiler performance and tibia ash. Overall, data suggest that the strategic utilization of enzyme combinations may provide additional benefits towards broiler production.

Mechanical and hand catching of broilers were performed to determine if differences existed in breast meat quality among catching methods. Meat quality results were summarized using a two-prong approach to investigate if average differences existed in meat quality through the determination of ultimate pH, rapid pH decline, color, water holding capacity, and tenderness within each treatment and by pinpointing individual quality problems within treatments. Two trials were conducted and revealed greater variation in meat quality during summer in comparison to winter within treatments in both catching methods. In winter, there was a lower incidence (p<0.05) of paleness in birds that were crated for 2h prior to euthanasia in comparison to birds immediately euthanized after catching within both hand and machine catching methods. Mechanical catching and crating for 2 hours was conducive to slightly better quality meat than hand catching in respect to averages and individual quality problems.

Over the past decade the poultry industry has made tremendous improvements in production and efficiency that can be attributed to areas of nutrition. Feed additives are one area of nutrition that the poultry industry utilizes to improve performance and efficiency. Experiment 1 investigated the effects of two nutrient densities diets and three enzyme inclusion strategies on 59 d Ross x Ross 708 male broiler performance under a natural disease challenge. These data suggest that when supplementing EIS2 broiler performance and processing characteristics are improved. Additionally, HD also improved broiler performance. The objective of the second experiment was to examine the interactive effects of probiotic and copper inclusion on 42 d Ross x Ross 708 male broiler performance. The resulting data suggests that inclusion of Cu-H at 250 ppm enhances broiler performance while lowering E.coli presence. These data also suggests that BL and Cu-H interact, ultimately reducing the efficacy of BL.

Three 2à 2 factorial experiments (EXP) were conducted to investigate the underlying mechanism of corn quality and a supplemented cocktail enzyme of amylase, protease, and xylanase on broiler performance from 0 to 49 days of age. In each of the first two EXP, the four diets used consisted of (1) diet AR-/BR-; reduced dietary energy by increasing corn A matrix metabolizable energy (ME) 138 kcal/kg in EXP 1 or 125 kcal/kg ME with corn B in EXP 2; (2) diet AR+/BR+; AR-/BR- plus Avizyme 1502® (AZ); (3) diet AN-/BN-; normal energy diet; (4) diet AN+/BN+; AN-/BN- plus AZ. In EXP 3, four dietary treatments came from EXP 1 and 2 consisted of: AN-, AN+, BN-, and BN+. For each EXP, 1,440 male Ross 708 chicks were randomly assigned to one of the four dietary treatments (trts) with 9 replicates (reps)/trt and 40 chicks/ rep on day (d) 0. Body weight gain (BWG) and feed intake (FI) were determined on feed change days (d 14, 28, and 37) and on d 7 and 49. On d 28, subsets of birds were transferred to Petersime batteries to evaluate feed passage rate and nutrient retention. Digesta samples from gizzard, jejunum, and ileum as well as pancreatic tissue samples were collected for analyses of amylase, protease, and xylanase activities. Intestinal tissue samples were collected for determination of maltase, sucrase, and aminopeptidase N activities. Jejunal tissue on d 28 was also collected for total RNA isolation and a subsequent genome-wide microarray assay. On d 50, 54 birds per trt were processed to evaluate carcass yield. Interactions of ME and AZ were observed in both EXP 1 and 2. In EXP 1, interactions of ME and AZ on BWG were observed with higher values in birds fed AN- while lower in birds fed AN+, while opposite observations in EXP 2 with corn B diets. In both EXP 1 and 2, feed conversion ratio (FCR) in birds fed normal energy diets was better (P <0.05) as compared to birds fed reduced energy diets after d 28. Percent fatpad was increased (P <0.05) with dietary energy levels in EXP 1. Percent pectoral minor was increased (P <0.05) in birds fed AZ diets (4.80 vs. 4.62%). In EXP 3, interaction of dietary corn and AZ (P < 0.05) on BWG was noted during d 14 and 37. Birds fed corn B diets had better (P < 0.05) BWG and feed efficiency as compared to birds fed corn A diets until d 14. Nitrogen retention on d 30 was greater (P < 0.05) in birds fed AN- and BN+ as compared to birds fed AN+ and BN-. A 2.8% more apparent metabolizable energy with nitrogen correction (AMEn) in corn A diets was observed as compared to corn B diets. Corn A had higher amylase and xylanase activities as compared to corn B. Xylanase activity in jejunal and ileal digesta of birds fed corn A diets were higher (P < 0.05) as compared to that of corn B birds on d 7 and 49. Sucrase-isomaltase contributed 63, 80, and 74 % of the total maltase activity in duodenum, jejunum, and ileum. Sucrase activities in duodenum and jejunum were correlated (P < 0.05) with performance, whereas duodenal aminopeptidase N was negatively correlated (P < 0.05) with performance except period BWG. Pancreatic amylase and protease as well as gizzard protease and xylanase activities were correlated (P < 0.05) with performance. Number of jejunal genes regulated (P < 0.05) by corn variety was 77 as compared to those by enzyme supplementation in corn A diets (30 genes) and corn B diets (23 genes). Immune response and metabolism related genes were the most regulated genes in birds fed different corn diets without enzyme addition. In conclusion, enzyme supplementation improved broiler performance. Dietary formulation strategy using either reduced energy or normal energy in associated with enzyme supplementation should base on the feed ingredient quality. Corn quality may come from active components such as protease inhibitor and xylanase, and improved performance in birds fed high quality corn diets might also relate with minimal immune response and metabolic demand. / Ph. D.

Growth and performance parameters of broiler chicks depend on adequate development of the small intestine. Stressors such as elevated or decreased temperatures during incubation and post-hatch transportation may have an effect on the gastrointestinal development of the broiler chick. The objective of the first study was to investigate the effects of elevated embryonic incubation temperature (IT) on post-hatch relative nutrient transporter gene expression, integrity of the intestinal epithelium, organ development, and performance in Ross 308 broiler chickens. Nine hundred fertile eggs were incubated at different egg-shell temperatures during development. Body weights and performance parameters were measured at day of hatch (DOH), d 7, 14, 21, 30, and 42. Small intestine and residual yolk were collected at DOH, d 2, 4, 6, and 10 and weighed individually. The small intestine was evaluated for mucosal morphology measurements and relative nutrient transporter (SGLT1, GLUT2, GLUT5, EAAT3, and PepT1) gene expression. The objective of the second study was to evaluate the effects of embryonic incubation, simulating a multi-stage incubation system, and post-hatch transportation temperatures on post-hatch performance, yolk free body weights, relative nutrient transporter gene expression, yolk utilization, intestinal morphology, and organ development of broiler chickens. Cobb 500 eggs (n=5200) were incubated with egg-shell temperatures, which were combined depending on the early and late development incubation periods as found in multi-stage incubators: Low (L): 36.7°C, Standard (S): 37.5°C, and High (H): 39°C. After hatch, chicks were further separated into 2 transportation groups: control (C; 34°C), and heat-stressed (D; 40°C). The eight resulting experimental groups were: LS-C, SS-C, LH-C, SH-C, LS-D, SS-D, LH-D, and SH-D. Three and two way interactions (P<0.05) were observed and discussed in both studies for all the parameters analyzed. These studies present for the first time the effects of altered embryonic IT and stress during transportation of newly hatched chicks, on small intestine morphology, digestive organ development, and expression of nutrient transporters mRNA in high-yield broiler chicks. These results contribute to the understanding of mechanisms by which either low or high temperatures, as compared to standard recommendations, during incubation and transportation can affect embryonic development and subsequent performance of broiler chicks. / Ph. D.

A series of experiments were conducted to evaluate the effects of dietary enzyme supplementation in broilers exposed to a live coccidia oocyst vaccine. In each experiment, Cobb 500 broilers were obtained at day of hatch from a commercial hatchery. Half of the chicks were sprayed with Coccivac BTM and housed in battery brooders in experiment (EXP) 1 and 2 or floor pens (EXP 3). Experimental diets were formulated according to Cobb 500 nutrient recommendations (positive control; PC) with the exception of Ca and available P (aP), which were reduced in the negative control (NC) diets approximately 0.1% (EXP 1), 0.11 and 0.13%, respectively (EXP 2), and 0.13% (EXP 3). Negative control diets in EXP 1 were supplemented with phytase (PHY), protease (PRO), xylanase (XYL), and the combination of PHY+PRO, PHY+XYL, and PHY+PRO+XYL. Negative control diets in EXP 2 were supplemented with PHY A, PHY B, and PHY C. In EXP 3, PC diets were supplemented with PHY at 1000 FTU/kg, and NC diets were supplemented with PHY at 1000 or 5000 FTU/kg. In all three experiments PHY supplementation generally improved (P ≤ 0.05) broiler performance and bone ash. Vaccination reduced (P ≤ 0.05) broiler performance in EXP 1 and 3, but increased (P ≤ 0.05) broiler performance in EXP 2. Xylanase and/or PHY supplementation tended to improve (P = 0.10) ileal amino acid digestibility (IAAD) in vaccinated broilers in EXP 1 and EXP 3. Phytase supplementation improved (P ≤ 0.05) IAAD and vaccination reduced (P ≤ 0.05) IAAD in EXP 3. Small intestinal morphology and goblet cell numbers were affected by enzyme supplementation and vaccination, which resulted in significant (P ≤ 0.05) interactions. In general, vaccination increased (P ≤ 0.05) small intestinal crypt depth and reduced (P ≤ 0.05) goblet cell numbers in EXP 3. Phytase supplementation of the NC diets fed to vaccinated broilers ameliorated the reduction in ileal goblet cells associated with vaccination (EXP 3). Enzyme supplementation tended to improve nutrient digestibility and altered small intestinal morphology in vaccinated broilers. Vaccination reduced broiler performance but nutrient digestion and dietary enzymes may improve nutrient utilization during a coccidia vaccination. / Ph. D.

Since the removal of in feed antibiotics in the past few years commercial poultry production is especially sensitive to the health of the small intestine. Healthy small intestines balance nutrient absorption and defensive barrier functions to ensure the chicken is able to meet the whole-body nutritional needs and is able to help prevent internalization of pathogens or potentially toxic components. This balance can only be maintained under stable conditions. When a disturbance event occurs the intestine imbalances until a new, and less efficient, balance can be achieved. The objective of this dissertation is to propose a novel model to understanding intestinal homeostasis in the face of various disturbance events. Chapter 2 investigated the effects of Runting Stunting Syndrome on broiler chickens in four different groups of chicks displaying clinical symptoms. The major finding in this study was that in two of the four groups the expression of stem cell gene Olfactomedin 4 was absent from the crypt though other functional genes were found to still be expressed there. Chapter 3 characterized intestinal gene expression following a single challenge of Eimeria acervulina in broiler chickens. During Eimeria infection gene expression of multiple host defense peptide genes were decreased compared to uninfected chickens. Further, Eimeria infected chickens increased cell proliferation within the crypt and post-peak infection showed signs of intestinal recovery. Additionally, chapter 3 developed a novel method for visualizing Eimeria as it infects the intestine. In chapters 4 and 5 cell type population changes during the peri-hatch intestinal maturation process were evaluated. Peri-hatch intestinal maturation is critical for the successful transition from embryonic to post-hatch life. Chapter 4 profiled changes in proliferative cells and gene expression of various stem cell marker genes during the peri-hatch period: the last three days of embryogenesis and the first week post-hatch. The stem cell marker gene Leucine Rich Repeat Containing G Protein-Coupled Receptor 5 (Lgr5) decreased during the post-hatch period while Olfactomedin 4 increased post-hatch. Both stem cell genes were expressed within the intestinal crypt, though prior to hatch Lgr5 was expressed in the lamina propria and villi as well. Additionally, the marker of proliferation Ki67 gene was expressed in cells throughout the intestine prior to hatch but became restricted to the crypts and along the center of the villi. Chapter 5 assessed the effect of providing probiotics to late term embryos via in ovo feeding (IOF). The effects of IOF were primarily observed on embryonic day 20 (e20), roughly 48 hours after IOF. On e20 the embryos in ovo fed probiotics in saline had increased expression in the ileum of Peptide Transporter 1 (PepT1) a marker gene for enterocytes and Mucin-2 (Muc2) a marker gene for goblet cells compared to non-injected control embryos. Also, on e20 the embryos in ovo fed saline only had numerically increased PepT1 and Muc2 compared to non-injected control embryos. The difference in responses between the probiotic and saline fed embryos on e20 suggests different routes of stimulation. These investigations illustrate various possible scenarios and means of investigating intestinal homeostasis during disturbance events. / Doctor of Philosophy / In healthy birds, the small intestine absorbs nutrients while preventing the free passage of microbes or toxic chemicals into the body. The two functions: absorption and barrier exclusion seem contradictory, but a balance is struck to ensure both functions continue. This balance-point, homeostasis, persists until an event disturbs it. Once disturbed the balance-point is changed and the intestine is unable to maintain both functions, until a new balance is found following recovery. The objective of the dissertation is to better understand intestinal homeostasis, through four different research projects. Experiment 1 characterized the intestinal cell population changes in broiler chickens during Runting Stunting Syndrome, a viral infection. The major finding of this chapter was that a stem cell gene, that is normally robustly expressed was not expressed in some groups of infected chicks but not all. Experiment 2 investigated the intestinal response of broiler chickens to Eimeria acervulina, an intestinal parasitic infection. Eimeria, which infects intestinal enterocytes, caused a decrease in defensive genes during the peak of infection. Then after the peak the intestine began to recover, as indicated by increased cell proliferation. Experiment 3 profiled changes in the expression patterns of stem cell and proliferation genes in the small intestine during the last days before hatch and the first week post-hatch. Pre-hatch stem and proliferative gene expression occurred in the crypt and villus, but became restricted to the crypt early during the post-hatch period. Experiment 4 assessed the effect of feeding probiotics to embryos before hatching on intestinal gene expression. Embryos fed probiotics had increased Mucin-2 and Peptide Transporter 1 gene expression in the last segment of the intestine, the ileum compared to non-fed embryos 48 hours after feeding. Additionally, treatments fed saline also showed increased gene expression, though to a lesser extent. Together these projects illustrate various disturbances to intestinal homeostasis and how intestinal cells change and respond during the disturbance and recovery periods.