Bobeck, Elizabeth

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Mitochondrial and Glycolytic Capacity of Peripheral Blood Mononuclear Cells Isolated From Diverse Poultry Genetic Lines: Optimization and Assessment

2022-01-28 , Lamont, Susan , Bobeck, Elizabeth , Bobeck, Elizabeth A. , Animal Science

Cellular metabolic preference is a culmination of environment, nutrition, genetics, and individual variation in poultry. The Seahorse XFe24 analyzer was used to generate foundational immune cellular metabolic data in layer, broiler, and legacy genetic strains using fresh chicken peripheral blood mononuclear cells (PBMCs). Baseline mitochondrial respiration [oxygen consumption rate (OCR)] and glycolytic activity [extracellular acidification rate (ECAR)] were determined in modern commercial laying hen (Bovans White) and broiler (Ross 308) lines, as well as the highly inbred lines of Iowa State University (L8, Fayoumi M-15.2, Spanish, Ghs-6), partially inbred broiler line, and advanced intercrosses of broiler by Fayoumi M-15.2 and broiler by Leghorn lines. Commercial broiler vs. Bovans layer and unvaccinated vs. vaccinated Bovans layer immune cell metabolic potential were compared following an in-assay pathway inhibitor challenge. Titrations consistently showed that optimal PBMC density in laying hens and broilers was 3 million cells per well monolayer. Assay media substrate titrations identified 25 mM glucose, 1 mM glutamine, and 1 mM sodium pyruvate as the optimal concentration for layer PBMCs. Pathway inhibitor injection titrations in Bovans layers and broilers showed that 0.5 μM carbonyl cyanide-4 phenylhydrazone (FCCP) and 1 μM oligomycin were optimal. Baseline OCR and ECAR were significantly affected by genetic line of bird (p < 0.05), with the dual-purpose, L8 inbred line showing the highest OCR (mean 680 pmol/min) and the partially inbred broiler line showing the greatest ECAR (mean 74 mpH/min). ECAR metabolic potential tended to be greater in modern layers than broilers (p < 0.10), indicating increased ability to utilize the glycolytic pathway to produce energy. OCR was significantly higher in vaccinated than unvaccinated hens (p < 0.05), while baseline ECAR values were significantly lower in vaccinated Bovans laying hens, showing increased oxidative capacity in activated immune cells. These baseline data indicate that different genetic strains of birds utilized the mitochondrial respiration pathway differently and that modern commercial lines may have reduced immune cell metabolic capacity compared with legacy lines due to intense selection for production traits. Furthermore, the Seahorse assay demonstrated the ability to detect differences in cellular metabolism between genetic lines and immune status of chickens.

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Algae-based feed ingredient protects intestinal health during Eimeria challenge and alters systemic immune responses with differential outcomes observed during acute feed restriction

2021-09-01 , Fries-Craft, K. , Meyer, M. M. , Bobeck, Elizabeth , Bobeck, Elizabeth , Animal Science

Compounds in microalgae-derived feed ingredients in poultry diets may improve intestinal physiology and immunity to protect against damage induced by physiological and pathogen challenges, but mechanisms are examined sparingly. The study objective was to evaluate changes to intestinal morphology, permeability, and systemic immunity in broilers fed a proprietary microalgae ingredient during 2 separate challenge studies. In study 1, two replicate 28 d battery cage trials used 200 Ross 308 broilers each (n = 400) fed a control diet ± 0.175% algae ingredient. Half of the birds were subjected to a 12 h feed restriction challenge and fluorescein isothiocyanate dextran (FITC-D) intestinal permeability assay on d 28. Study 2 used 800 broilers randomly assigned to the same dietary treatments and housed in floor pens for 42 d. At d 14, intestine and spleen samples were collected from 10 birds/ diet. Half of the remainder was orally inoculated with 10X Coccivac-B52 vaccine in a 2 × 2 factorial treatment design (diet and Eimeria inoculation). The FITC-D assay was conducted at 1, 3, 7, and 14 d post-inoculation (pi) while intestinal and spleen samples were collected at 3, 7, 14, and 28 dpi for histomorphology and flow cytometric immune cell assessment. Study 1 validated intestinal leakage via FITC-D absorbance induced by feed restriction but showed no algae-associated protective effects. In study 2, algae preserved intestinal integrity during coccidiosis (P = 0.04) and simultaneously protected jejunal villus height as early as 7dpi (P < 0.0001), whereas intestinal damage resolution in control birds did not occur until 14 dpi. Algae inclusion increased splenic T cells in unchallenged broilers at d 14 by 29.6% vs. control (P < 0.0001), specifically γδ T cell populations, without impacting performance (P < 0.03). During Eimeria challenge, splenic T cells in algae-fed birds did not show evidence of recruitment to peripheral tissues, while control birds showed a 16.7% reduction compared to their uninoculated counterparts from 3 to 7 dpi (P < 0.0001). This evidence suggests the algae ingredient altered the immune response in a manner that reduced recruitment from secondary lymphoid organs in addition to protecting intestinal physiology.

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Growth performance of male broilers fed ExPress® soybean meal and high-shear dry extruded corn

2021-01-01 , Meyer, Meaghan , Bobeck, Elizabeth , Bobeck, Elizabeth , Animal Science

Soybean meal (SBM), a co-product of soybean oil obtained via solvent or mechanical extrusion, is the key protein source in broiler diet formulations. Mechanical extrusion, or high-temperature processing with high-shear dry extrusion, has demonstrated 1 to 2% increased amino acid digestibility in broilers versus solvent-extracted SBM through reduction of antinutritional factors (ANF). High-shear dry extrusion followed by mechanical pressing using a screw press (extruding-pressing) produces soy oil and partially de-oiled soymeal (ExPress). The extruding-pressing process has been growing in popularity due to improved cost and material efficiency by eliminating the need for solvents or steam dryers in the production of animal feed ingredients. Corn extrusion is likewise linked with increased nutritive value through improved palatability and approximately 20% increased starch gelatinization, resulting in superior growth performance and in one reported case, reduced mortality in broilers. In the present study, high-shear dry extruded corn and ExPress SBM were formulated in combination with conventional corn and solvent-extracted SBM to determine impact on growth performance in male broilers. Extruded corn increased feed intake and ExPress SBM increased weight gain. The diet containing dry extruded corn and ExPress SBM outperformed all other diet combinations, including unprocessed, ground corn and conventional, solventextracted SBM variations. The diet containing unprocessed, ground corn and ExPress SBM had the most efficient overall feed conversion ratio (FCR). The combination of lower feed intake observed with the ground corn ingredient and the improved protein digestibility reflected by superior growth in ExPress SBM made this the ideal formulation, out of the 4 diets examined, in terms of feed efficiency.

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Laser Environmental Enrichment and Spirulina Algae Improve Broiler Growth Performance and Alter Myogenic Gene Expression and pectoralis major Dimensions

2021-12-14 , Johnson, Anna , Bobeck, Elizabeth , Bobeck, Elizabeth A. , Animal Science

Sustainability in poultry production is evident in efforts to reduce inputs and a focus on bird welfare and livability. Dietary protein alternatives to traditional sources such as soybean meal aim to meet or exceed efficiency benchmarks and be cost-effective. Environmental enrichment encouraging activity may reduce the occurrence of the predominant breast muscle myopathy, woody breast (WB); interventions to minimize muscle damage and economic loss have yet to be established. The study objectives were to maintain or improve broiler performance and breast quality through environmental enrichment and partially replacing dietary soybean meal with Spirulina. Twelve hundred Ross 708 broilers were randomly assigned to enrichment (LASER; laser enrichment, or CON; no laser enrichment) and diet (algae; 2.5% Spirulina algae, or control) in a 2 × 2 factorial design for 49 days. The same 70 randomly selected birds were examined for contact dermatitis wk 1–6. Breast width was measured weekly on 200 growing broilers beginning on d22. On d42 and 49 slaughter, WB score was assigned using a tactile 0–3 scale and the right breast filet was weighed (n = 200). RNA isolated from 30 breast muscle samples each at d42 and 49 was analyzed using real-time qPCR. Laser enrichment increased body weight at all timepoints (d49: 0.148 kg, P < 0.001). Feed conversion ratio was improved in LASER-enriched birds by 3 points in the starter period (P = 0.003). Breast width was increased at all timepoints in LASER-enriched birds compared to CON (d49: 0.47 cm, P < 0.001). Algae inclusion increased body weight at d28 (0.059 kg, P = 0.005). At d42, 12% more LASER-enriched WB scores were 0 (normal) compared to CON, and at d49, 15% more enriched scores were 0. At d42, 5% more algae-fed broiler scores were 0 compared to control. LASER-enriched broiler breast tissue showed upregulated expression of myogenin, muscle regulatory factor 4, insulin-like growth factor 1, and myostatin compared to CON (P < 0.01). Both laser enrichment and algae inclusion improved broiler performance without negatively impacting environmental or physiological outcomes. LASER enrichment decreased severity of WB score and positively shifted myogenic gene expression in the breast muscle at slaughter.

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Comparison of Naturally Occurring vs. Experimental Infection of Staphylococcus aureus Septicemia in Laying Hens in Two Different Age Groups

2021-06-01 , Meyer, Meaghan , Bobeck, Elizabeth , Sato, Yuko , Bobeck, Elizabeth , El-Gazzar, Mohamed , Animal Science , Veterinary Diagnostic and Production Animal Medicine

In April and November of 2018, multiple commercial laying hen flocks within the same company presented with a sharp increase in mortality and drop in egg production that persisted for several days. These flocks showed striking necropsy lesions consistent with systemic infection and responded to antimicrobial treatment in the feed. Staphylococcus aureus (SA) was the most frequently isolated organism from multiple tissues including comb and wattle lesions, lungs, liver, ovary, spleen, and bone marrow. Given such an uncommon presentation of SA, which is known as a secondary opportunistic pathogen, a challenge study was conducted to evaluate its role in these disease outbreaks. In the present study, laying hens of two ages (22 and 96 wk) were inoculated with SA via three routes: oral gavage, subcutaneous (SC) injection, and intravenous (IV) injection. Both young and old hens in the IV group showed a significant increase in body temperature and drop in body weight; however, the clinical signs observed in the naturally occurring outbreaks were not present. SA was reisolated at multiple time points postchallenge from all challenge groups except the negative control group. While the SC group showed localized necrosis at the injection site, microscopic changes were different from changes observed in birds from the natural outbreaks. Despite observed initial differences in route and age, the SA challenge strain was not capable of reproducing the disease on its own. The results of this study indicate that SA may have played a role in the increased mortality, clinical signs, and necropsy lesions reported with the naturally occurring outbreaks. However, SA should still be considered as a secondary opportunistic pathogen. Other factors that could have caused the initial insult are stress, immunosuppression, or other primary infectious agents. The results of this study may aid veterinary diagnosticians, clinicians, and all poultry professionals to include SA in their differentials list as a secondary opportunistic pathogen in similar cases. This is an uncommon presentation and further field observations and clinical studies are needed to better elucidate the pathogenesis of this disease, which will in turn help to prevent future outbreaks.

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Laser Enrichment Device Stimulates Broiler Laser-Following Behavior While Increasing Individual Bird Locomotion and Pen-Wide Movement

2021-11-29 , Johnson, Anna , Bobeck, Elizabeth , Bobeck, Elizabeth A. , Animal Science

Genetic selection for fast growth rate and high breast muscle yield in modern broilers has unintended effects on animal welfare and behavior, namely in terms of inactivity and leg disorders. We hypothesized that exercise stimulated through environmental enrichment could positively stimulate pen-wide activity and improve bird welfare. The study objectives were to implement a laser enrichment device to motivate active and feeding behaviors throughout the pen. Twelve hundred Ross 708 broilers were randomly assigned to enrichment (LASER; laser enrichment, or CON; no laser enrichment) for 49 d. Seventy focal birds were randomly assigned to 14 video-recorded pens for behavioral analysis, including focal bird home pen behavior and walking distance. Pen-wide activity was also measured during the 4-min laser periods, four times daily, d0–8, and 1 day weekly, wk 1–6. Focal birds were gait scored wk 1–6, and were euthanized on d42 for tibia bone mineral content, density, and bone breaking strength analysis. Time spent active was increased in LASER-enriched birds compared to CON on wk 3–5 by up to 214% (wk 4), and percent of time at the feeder was increased in LASER-enriched birds by 761% on wk 4 (P < 0.05). Peak percent of birds following the laser (LASER-enriched pens only) was observed on d0 (8.52%). Over wk 1–6, peak laser-following behavior was observed on wk 3 (3.07% of birds). Percent of birds moving during laser periods was increased in LASER-enriched pens on d0, 1, 2, 6, 7, and 8, with a percent increase of 68.7% observed on d1 (P < 0.05). Percent of birds moving (laser-following or not) was increased on wk 1, 3, and 4 in LASER-enriched pens, with an increase of 69.7% observed on wk 4 (P < 0.05). No differences were found in tibia measures. These data indicate that laser enrichment stimulated voluntary locomotion through wk 5 and laser-following behavior through wk 6, and that the relatively small percent of birds actively following the laser stimulated pen-wide movement above the level of the CON through wk 4 on study.

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Age and Staphylococcus aureus Inoculation Route Differentially Alter Metabolic Potential and Immune Cell Populations in Laying Hens

2021-03 , Fries-Craft, Krysten , Meyer, Meaghan M , Sato, Yuko , El-Gazzar, Mohamed , Bobeck, Elizabeth A. , Animal Science , Veterinary Diagnostic and Production Animal Medicine

In 2018 and 2019, Staphylococcus aureus was isolated from multiple post-molt commercial laying hens with unusually high mortality. A challenge study was conducted to elucidate the role of S. aureus in this disease outbreak and the work herein represents the assessment of immunological responses in laying hens experimentally infected with S. aureus isolates from these cases. A total of 200 laying hens at 22 or 96 weeks of age (100/ age group) were assigned to 1 of 4 experimental inoculation groups (negative control, oral gavage, subcutaneous injection, or intravenous injection) after a 72 h acclimation period. Blood samples were taken prior to inoculation (baseline), 6 h post-inoculation (pi), 24 hpi, 3 dpi, and 7 dpi. Additional spleen samples to further assess systemic immunity were taken at baseline, 3 and 8 dpi. Metabolic phenotypes of peripheral blood mononuclear cells (PBMC) were isolated and assessed by Seahorse metabolic assay. Immune cell profiles in the spleen and PBMC were assessed by multicolor flow cytometry. At baseline, 96-week-old laying hens had 26.7% fewer PBMC-derived T cells compared to 22-week-old birds. Older hens had 28.9% increased helper T cell (TH) populations and 60.5% reduced γδ T cells (P = 0.03 and < 0.0001) which may contribute to variable clinical responses between age groups; however, no age-related differences in metabolic potential were observed. Metabolic outcomes showed that birds remained stressed from transport and re-housing past a 72 h acclimation period and through 24 h- 3 days post-inoculation. Inoculation with S. aureus generally reduced oxidative and glycolytic potentials compared to the control, with the greatest reductions observed in birds inoculated by intravenous injection (P < 0.05). Overall CD3+ T cell populations showed significant reductions in the intravenous group compared to other inoculation routes from 24 hpi to 7 dpi (23.6–39.0%; P ≤ 0.0001). These results suggest that age-related baseline differences in T cell populations and changes to T cell subpopulations and other immune cells due to inoculation route may have an additive effect on S. aureus- induced reductions in metabolic potential; however, further research linking metabolic potential and immune cell profiles is needed.