Graded levels of Eimeria infection linearly reduced the growth performance, altered the intestinal health, and delayed the onset of egg production of Hy-Line W-36 laying hens when infected at the prelay stage.

The aim of this experiment was to investigate how different levels of Eimeria infection affect the performance, intestinal health, oxidative status, and egg production of Hy-Line W-36 pullets and laying hens. Three hundred and sixty Hy-Line W-36 pullets, aged 15 wk, were randomly distributed into 5 treatment groups, each comprising 6 replicates and a nonchallenged control. At 15 wk, pullets were inoculated with different levels of mixed Eimeria species as high-dose, medium-high, medium-low, and low-dose treatments. The growth performance and average daily feed intake (ADFI) were measured from 0- to 18-days postinoculation (DPI), whereas hen day egg production (HDEP) was recorded from wk 19. The markers of gastrointestinal health and oxidative status were measured at 6 DPI, 14 DPI, and 23 wk of age. The findings revealed a significant linear reduction in growth performance in response to increased Eimeria challenge dosage on 6 and 14 DPI (P < 0.0001, P-L < 0.0001). An interaction between the graded level of Eimeria infection and DPI was observed for ADFI. The challenged pullets showed a reduction in ADFI starting at 4 DPI, which persisted until 14 DPI, when ADFI recovered back to normal. The most significant drop in feed intake was observed in 6 DPI in all the Eimeria-infected groups. The markers of gastrointestinal health (gastrointestinal permeability and tight junction proteins) were upregulated in challenged pullets because of infection, whereas the relative mRNA expression of key nutrient transporters was downregulated following infection on 6 and 14 DPI (P < 0.05). As a result of an infection on 6 DPI, the oxidative equilibrium was shifted toward the oxidative stress, and at the same time, upregulation of proinflammatory and inflammatory cytokines was observed (P < 0.05). An interaction between the Eimeria challenge dosage and bird age was observed for HDEP (P = 0.0427). The pullets infected with Eimeria started to lay eggs later than the Control birds. However, the HDEP of the challenged groups became similar to Control only at wk 22, 3 wk after laying eggs. In conclusion, coccidiosis reduced growth performance, altered gastrointestinal health, induced oxidative stress, and delayed egg production when infected at the prelay stage of pullets and negatively impacted the laying hens' overall performance.

Effect of the combination of 25-hydroxyvitamin D3 and higher level of calcium and phosphorus in the diets on bone 3D structural development in pullets.

Bone issues such as osteoporosis are major concerns for the laying hen industry. A study was conducted to improve bone-health in pullets. A total of 448 one-day-old Hyline W36 pullets were randomly assigned to four treatments (8 rep; 14 birds/rep) until 17 weeks (wks). Dietary treatments were: 1) vitamin D3 at (2,760 IU/kg) (D), 2) vitamin D3 (2,760 IU/kg)+62.5 mg 25-(OH)D3/ton (H25D), 3) vitamin D3 (2,760 IU/kg) + 62.5 mg 25-(OH)D3/ton + high Ca&P (H25D + Ca/P), and 4) vitamin D3 (2,760 IU/kg) + high Ca&P (D + Ca/P). The high calcium (Ca) and phosphorus (P) diet was modified by increasing both high calcium and phosphorus by 30% (2:1) for the first 12 wks and then only increasing P for 12-17 wks to reduce the Ca to P ratio. At 17 wk, growth performance was measured, whole body composition was measured by dual energy x-ray absorptiometry (DEXA), and femur bones were scanned using Micro-computed tomography (Micro-CT) for bone 3D structure analyses. The data were subjected to a one-way ANOVA using the GLM procedure, with means deemed significant at p < 0.05. There was no significant outcome for growth performance or dual energy x-ray absorptiometry parameters. Micro-computed tomography results indicated that the H25D + Ca/P treatment had lower open pore volume space, open porosity, total volume of pore space, and total porosity in the cortical bone compared to the D + Ca/P. It also showed that a higher cortical bone volume/tissue volume (BV/TV) in the H25D + Ca/P than in the D + Ca/P. Furthermore, the H25D + Ca/P treatment had the lowest trabecular pattern factor and structure model index compared to the other treatments, which indicates its beneficial effects on trabecular structural development. Moreover, the H25D + Ca/P had a higher trabecular percentage compared to the D and 25D, which suggests the additional high calcium and phosphorus supplementation on top of 25D increased trabecular content in the cavity. In conclusion, the combination of 25D with higher levels of high calcium and phosphorus could improve cortical bone quality in pullets and showed a beneficial effect on trabecular bone 3D structural development. Thus, combination of a higher bio-active form of vitamin D3 and higher levels of high calcium and phosphorus could become a potential feeding strategy to improve bone structural integrity and health in pullets.

Graded levels of Eimeria challenge altered the microstructural architecture and reduced the cortical bone growth of femur of Hy-Line W-36 pullets at early stage of growth (0-6 wk of age).

An experiment was carried out to evaluate the impact of mixed Eimeria challenge on skeletal health of Hy-Line W-36 pullets. A total of 540, 16-day-old pullets were randomly allocated into 5 treatment groups, including a nonchallenged control. A mixed Eimeria species solution containing 50,000 E. maxima, 50,000 E. tenella, and 250,000 E. acervulina oocysts per mL was prepared and challenged to 1 group as a high-dose treatment. The 2-fold serial dilution was done to prepare the medium-high (25,000 E. maxima; 25,000 E. tenella; 125,000 E. acervulina), the medium-low (12,500 E. maxima; 12,500 E. tenella; 62,500 E. acervulina), and the low (6,250 E. maxima; 6,250 E. tenella; 31,250 E. acervulina) dose treatments which were challenged to 3 corresponding groups, respectively. The mineral apposition rate (MAR) was measured from 0 to 14 d post inoculation (DPI) and 14 to 28 DPI using calcein injection. The microstructural architecture of the femur was analyzed using the Skyscan X-ray microtomography (microCT) on 6, 14, and 28 DPI. The results showed that the MAR decreased linearly with an increase in the challenged dose (P < 0.05) during 0 to 14 DPI. The results of microCT revealed that cortical and total BMD, BMC, bone volume (BV), and bone volume as a fraction of tissue volume (BV/TV) of femur decreased both linearly (P < 0.05). Conversely, the total number of pores increased linearly with an increase in challenge dosages on 6 and 14 DPI. Trabecular BMD, BV, BV/TV, trabecular number, and trabecular thickness decreased linearly with an increase in the challenge dosages (P < 0.05) on 6 DPI. Furthermore, Eimeria infection significantly increased the number of osteoclasts and osteoclastic activity (P = 0.001). The result of this study suggests that the mixed Eimeria challenge negatively impacts the quality of skeletal health in a linear or quadratic manner with an increase in the concentration of Eimeria oocysts. The negative impact on long bone development might be due to malabsorption, nutrient deficiency during the infection, along with oxidative stress/inflammation disrupting the balance of osteoblastic and osteoclastic cells and their functions.

Impacts of phytase and coccidial vaccine on growth performance, nutrient digestibility, bone development, and intestinal gene expression of broilers fed a nutrient reduced diet.

An experiment was conducted to evaluate effects of phytase and coccidial vaccine on growth performance, bone ash, bone 3-D microstructure, nutrient digestibility, and gene expression of intestinal biomarkers in broilers fed a regular or nutrient-reduced diet. The experiment was conducted in a 2 × 4 factorial arrangement with 6 replicates per treatment and 10 birds per replicate. Two main factors were coccidial vaccine and dietary treatments. The dietary treatments included: 1) a positive control (PC; normal nutrient levels); 2) a negative control (NC; with a reduction of 0.15% of Ca and avP and 5% of essential amino acid (EAA) and crude protein relative to PC); 3) NC + 500 FTU/kg of phytase; and 4) NC + 1,500 FTU/kg of phytase. No interaction effect of phytase and coccidial vaccine on growth performance, bone ash, and apparent ileal digestibility (AID) was observed. For the main effect, birds fed the NC diet showed lower (P = 0.007) BWG during d 0 to 21 compared to PC birds, whereas supplementing 500 or 1,500 FTU/kg phytase increased BWG to the similar level to the PC. During d 0 to 21, vaccinated birds had a lower (P < 0.001) FI and better (P = 0.045) FCR compared to unvaccinated birds. Birds fed the NC diet resulted a decrease in tibia fat-free dry bone weight (P = 0.012), ash weight (P = 0.005), ash percentage (P < 0.001), and ash concentration (P = 0.019) compared to the PC group at d 21, whereas supplementing phytase at 500 or 1,500 FTU/kg in NC diet was able to improve these bone parameters to the similar level to the PC; however, vaccination did not have any effect on bone ash. Similarly, birds fed with the NC diet showed had significant lower bone microstructure levels including bone volume, bone mineral density, and bone mineral content (P < 0.001), and supplementing phytase at 1,500 FTU/kg improved these parameters. Vaccination improved AID of nitrogen (P < 0.001). Birds from the NC and both phytase supplementation groups had a higher (P = 0.001) AID of Ca compared to the PC. Supplementing phytase at 500 FTU/kg or 1,500 FTU/kg improved (P < 0.001) AID of P compared to the NC. Additionally, the NC had a lower AID of DM than the PC, whereas supplementing phytase at 500 FTU/kg or 1,500 FTU/kg improved DM digestibility (P = 0.0299). In conclusion, supplementation of phytase at 500 or 1,500 FTU/kg improved growth performance, bone mineralization, and nutrient digestibility regardless of vaccination, with a more pronounced effect when supplementing phytase at 1,500 FTU/kg.

Partial Replacement of Soybean Meal with Canola Meal or Corn DDGS in Low-Protein Diets Supplemented with Crystalline Amino Acids-Effect on Growth Performance, Whole-Body Composition, and Litter Characteristics.

A 42-day study was conducted to explore the application of supplemental amino acids (AA) in low-protein diets with soybean meal (SBM), canola meal (CM) or corn distillers dried grain with solubles (cDDGS) as the main protein feedstuffs. The responses of interest were growth performance, carcass yield, whole-body composition, litter ammonia and litter N. On d 0, a total of 540 Cobb 500 (off-sex) male broilers were allocated to 36 floor pens. All the birds received one starter diet that met nutrient requirements during the first 10d. Thereafter, six experimental diets were provided in grower and finisher phases. The diets included a positive control (PC): a corn−SBM diet with adequate protein. The protein level of the negative control (NC) was decreased by 45 g/kg relative to the PC. The next two diets had the same protein levels as the NC but with cDDGS added at 50 or 125 g/kg. The last two diets had the same CP as the NC but with CM added at 50 or 100 g/kg. All the low-protein diets had the same level of standardized ileal digestible indispensable AA according to Cobb 500 recommended level. Gly and Ser were added as sources of non-specific N. The dietary protein reduction in corn−SBM diets at both phases decreased (p < 0.05) weight gain and increased (p < 0.05) feed conversion ratio (FCR). Increasing levels of cDDGS or CM, at a constant CP level, linearly decreased (p < 0.05) the weight gain and feed intake, whereas increasing CM level linearly increased (p < 0.05) FCR in the grower and finisher phases. The eviscerated and carcass yields decreased, whereas the fat yield increased (p < 0.05) with reduced protein in corn−SBM diet. Increasing levels of cDDGS and CM at a constant CP level quadratically decreased (p < 0.05) the eviscerated weight, whereas the fat weight linearly decreased (p < 0.05) with increasing levels of cDDGS and CM. The birds receiving the PC diet had a lower (p < 0.05) lean muscle (%) and a higher fat (%) compared to birds receiving the NC diet at d 21. However, on d42, birds receiving the PC diet had decreased (p < 0.05) bone mineral density, bone mineral content and lean weight compared to those receiving the NC diet. The litter ammonia increased (p < 0.05) with the increasing levels of protein in the SBM diets. In conclusion, 50 g/kg inclusion levels of CM and cDDGS at the same low-protein levels as SBM produced a similar growth response to the NC, whereas higher levels were detrimental. Hence under the conditions of the current experiment, complete replacement of SBM with DDGS or CM in low-protein diets was not feasible.

Effects of mixed Eimeria challenge on performance, body composition, intestinal health, and expression of nutrient transporter genes of Hy-Line W-36 pullets (0-6 wks of age).

A study was aimed to investigate the effects of mixed Eimeria challenge on performance, gastrointestinal health, oxidative stress, inflammation, and expression of nutrient transporter genes of Hy-Line W-36 pullets. A total of 540, 16-d old pullets were randomly allocated into 5 treatment groups with 6 replicate cages, including a nonchallenged control group. A mixed Eimeria species solution containing 50,000 E. maxima, 50,000 E. tenella, and 250,000 E. acervulina oocysts per mL was prepared and challenged to one group as a high-dose treatment (High). The 2-fold serial dilution was done to prepare the medium-high (Med-High: 25,000 E. maxima; 25,000 E. tenella; and 125,000 E. acervulina), the medium-low (Med-Low: 12,500 E. maxima; 12,500 E. tenella; and 62,500 E. acervulina), and the low (Low: 6,250 E. maxima; 6,250 E. tenella; and 31,250 E. acervulina) dose treatments, and these dosages were challenged to 3 remaining groups, respectively. Growth performance, daily feed intake (FI), and mortality were calculated from 0-14 d postinfection (DPI). Gastrointestinal permeability (GP) was measured on 3, 5, 6, 7, and 9 DPI. The result indicated significant linear responses to the Eimeria challenge dosage in average body weight and body weight gain (P < 0.0001). An interaction between treatment and DPI was observed for FI (P < 0.0001). Feed intake significantly dropped from 4 DPI and did not recover until 12 DPI in the challenged groups. The lowest FI for each of the challenged groups was observed on 5 DPI. Gastrointestinal permeability increased linearly, peaking at 5 DPI, and was recovered back to normal by 9 DPI in the challenged groups. Furthermore, gene expression of tight junction proteins was linearly upregulated by increased Eimeria dosages. The oxidative status of the pullets was lowered in the challenged groups than the nonchallenged control group, whereas the expression of inflammatory and proinflammatory cytokines was upregulated by Eimeria challenge on 6 DPI (P < 0.05). The highest mortality was observed in pullets challenged with the High, followed by the Med-High (P < 0.0001) on 5 DPI. In summary, the mixed Eimeria challenge linearly reduced the growth performance of pullets with an increase in oxidative stress and inflammation. A severe effect of Eimeria on gastrointestinal health was observed on 5 or 6 DPI as suggested by GP, tight junction genes, and mortality results. This study indicates that Eimeria infection can be a threat to gastrointestinal health related issues in pullets.

Effect of Dietary Supplementation of Probiotic Aspergillus niger on Performance and Cecal Microbiota in Hy-Line W-36 Laying Hens.

This study aimed to investigate the role of the probiotic Aspergillus niger on the production performance, egg quality, and cecal microbial load of Clostridium perfringens, Salmonella spp., and Escherichia coli in Hy-Line W-36 laying hens. A total of 72, 45-week-old Hy-Line W-36 laying hens were randomly allocated to one of the three dietary treatments with six replicates, and each replicate had four individually caged laying hens (n = 6 and 4 hens/replicate). The hens in each treatment group were fed a corn and soybean meal diet (Control), a diet supplemented with bacitracin methylene disalicylate (BMD) at a rate of 495 mg/kg of feed (Positive Control), or a diet supplemented with Aspergillus niger (Probioist®) at a rate of 220 mg/kg of feed (Probiotic). Supplementing probiotics in the laying hen diet significantly increased egg production at weeks 3 and 6 compared with the Positive Control. Haugh unit, a measure of egg quality, was significantly higher in laying hens fed the probiotic diet compared with the Control or Positive Control at week 10. Furthermore, the Probiotic group had numerically lower cecal microbial loads of pathogenic bacteria (Clostridium perfringens, Salmonella spp., and Escherichia coli) compared with the Control and Positive Control groups. The results suggest that Aspergillus niger could be used as a probiotic to improve laying hen performance and egg quality.

Role of 1,25-Dihydroxyvitamin D3 on Osteogenic Differentiation and Mineralization of Chicken Mesenchymal Stem Cells.

1,25-dihydroxyvitamin D3 (1,25OHD) has been suggested to play an important role in osteogenic differentiation and mineralization. However, limited data have been reported in avian species. In the present study, the direct role of 1,25OHD on osteogenic differentiation and mineralization in chicken mesenchymal stem cells (cMSCs) derived from day-old broiler bones was investigated. cMSCs were treated with control media (C), osteogenesis media (OM), OM with 1, 5, 10, and 50 nM 1,25OHD, respectively. The messenger RNA (mRNA) samples were obtained at 24 and 48 h and 3 and 7 days to examine mRNA expression of key osteogenic genes [runt related transcription factor 2 (RUNX2), bone morphogenetic protein 2 (BMP2), collagen type I alpha 2 chain (COL1A2), bone gamma-carboxyglutamate protein (BGLAP), secreted phosphoprotein 1 (SPP1), and alkaline phosphatase (ALP)]. Cells were stained at 7, 14, and 21 days using Von Kossa (mineralization), Alizarin Red (AR; mineralization), and Alkaline Phosphatase (early marker) staining methods. From the mRNA expression results, we found a time-dependent manner of 1,25OHD on osteoblast differentiation and mineralization. In general, it showed an inhibitory effect on differentiation and mineralization during the early stage (24 and 48 h), and a stimulatory effect during the late cell stage (3 and 7 days). The staining showed 1,25OHD had an inhibitory effect on ALP enzyme activities and mineralization in a dosage-dependent manner up to 14 days. However, at 21 days, there was no difference between the treatments. This study provides a novel understanding of the effects of 1,25OHD on osteogenic differentiation and mineralization of cMSCs depending on cell stage and maturity.

Role of 25-Hydroxyvitamin D3 and 1,25-Dihydroxyvitamin D3 in Chicken Embryo Osteogenesis, Adipogenesis, Myogenesis, and Vitamin D3 Metabolism.

A study was conducted to understand the effects of 25-hydroxyvitamin D3 (25OHD) and 1,25-dihydroxyvitamin D3 (1,25OHD) administration on the expression of key genes related to osteogenesis, adipogenesis, myogenesis, and vitamin D3 metabolism in the chicken embryo. A total of 120 fertilized Cobb 500 eggs were used in the current study and were reared under standard incubation conditions. On embryonic day 3 (ED 3), PBS (C), PBS with 40ng 1,25OHD (1,25D-L), 200ng 1,25OHD (1,25D-H), 40ng 25OHD (25D-L), or 200ng 25OHD (25D-H) were injected into the dorsal vein of developing embryos. Whole embryos were harvested at 1, 3, and 6h post-injection for gene expression analyses (n=8). Gene expression for key osteogenesis markers (RUNX2: runt-related transcription factor 2; BMP2: bone morphogenetic protein 2; COL1A2: collagen type I alpha 2 chain; BGLAP: bone gamma-carboxyglutamate protein; SPP1: secreted phosphoprotein 1; and ALP: alkaline phosphatese), adipogenesis markers (PPAR-γ: peroxisome proliferator-activated receptor gamma; FASN: fatty acid synthase; and FABP4: fatty acid binding protein 4), myogenesis markers (MYOG: myogenin; MYOD1: myogenic differentiation 1; and MYF5: myogenic factor 5), and the enzyme responsible for vitamin D3 inactivation (CYP24A1: cytochrome P450 family 24 subfamily A member 1) were measured using real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). Data were normalized by the ΔΔCT method and analyzed using a one-way ANOVA. Results indicated that at 1h post-injection, no differences were found among treatments. At 3h, the early osteogenesis differentiation marker, ALP, was increased by 1,25D-H and 25D-H, and 25D-H also stimulated the expression of adipogenesis markers (FAPB4 and FASN). In contrast, the expression of myogenesis markers (MYOD1 and MYF5) was suppressed by 25OHD or 1,25OHD treatments, respectively. At 6h, a late osteogenic differentiation marker, SPP1, was increased by 25D-H. MYOD1 and MYF5 were continuously suppressed by 25OHD treatments or 1,25D-H. The evidence of vitamin D3 metabolite retention was assessed by measuring CYP24A1 expression. At 1h, there were no differences in CYP24A1 expression. At 3h, all treatments upregulated CYP24A1 expression relative to control (PBS) embryos. However, at 6h, only the 25D-H group retained higher CYP24A1 expression compared to the other treatments. In conclusion, the results suggested both 1,25OHD and 25OHD induced chicken embryo osteogenesis and adipogenesis, but inhibited myogenesis during early chicken embryo development. The higher dosage of 25OHD showed a possibility of a longer retention time in the embryos.

Effects of dietary protein, energy and ß-mannanase on laying performance, egg quality, and ileal amino acid digestibility in laying hens.

ß-mannan is a nonstarch polysaccharide found in hulled and dehulled soybeans that can survive drying-toasting phase of processing soybeans and have antinutritive effects in poultry. ß-mannanase is an active enzyme (endohydrolase) that can hydrolyze ß-mannan to reduce its antinutritional effects. Two experiments were conducted to evaluate the effects of ß-mannanase supplementation in low energy/protein diets on egg production, egg quality, and apparent ileal digestibility of the dry matter (DM), crude protein (CP), and amino acids in 21-week-old Single Comb White Leghorn hens (Hy-Line W-36). A total of 192 hens (8 replicates of 6 hens per treatment) for a production study (Exp. 1) and a total of 64 hens (8 replicates of 2 hens per treatment) for a digestibility study (Exp. 2) were randomly allocated to 4 experimental treatments in a 2 × 2 factorial arrangement. Four dietary treatments were control (CS) based on corn and 44% CP soybean meal (ME: 2,850 kcal/kg CP: 18.5%) and CS-low energy/protein (CSL) (ME: 2,750 kcal/kg CP: 17.5%), with or without 0.05% ß-mannanase enzyme. Hens were fed the experimental diets for 14 d for the digestibility study and 8 wk for the production study. Hen-day egg production (HDEP), weekly feed intake, FCR, and biweekly egg quality parameters were measured. Significant interaction on feed intake (P < 0.01) was observed between energy/protein and enzyme. At 3, 6 and 8 wk, the feed intake and FCR of CSL with enzyme were significantly lower than those of CSL without enzymes. The main effects indicated that birds fed diets without inclusion of ß-mannanase had higher feed intake than those fed diets with enzymes at 4, 7, and 8 wk. The inclusion of ß-mannanase significantly increased (P < 0.05) HDEP at 2, 3, 5, and 7 wk. However, there was no significant effect of nutrient density or enzyme supplementation on egg quality parameters. The digestibility study showed that the inclusion of ß-mannanase significantly improved (P < 0.01) apparent ileal digestibility of lysine, histidine and tryptophan in the diet. The results of these experiments indicate that supplementation of ß-mannanase could reduce the feed intake and FCR and improve HDEP and apparent ileal digestibility of key amino acids in corn/soy diets fed to laying hens.

Effects of the housing environment and laying hen strain on tibia and femur bone properties of different laying phases of Hy-Line hens.

This study aimed to determine the effect of the housing environment and laying hen strain on tibia and femur properties. A 3 × 2 factorial arrangement of 3 housing environments (conventional cages [CC], enriched colony cages [EC], and free range [FR]) and 2 laying hen strains (Hy-Line W-36 [W-36] and Hy-Line Brown [HB]) in a completely randomized design was conducted from 32 to 85 wk of age. Six left tibias were collected at 8 different time points (38, 45, 52, 59, 65, 72, 79, and 85 wk of age), whereas 6 left femurs were collected at 3 time points (38, 65, and 85 wk of age). Tibias were evaluated for tibia breaking strength (TBS) and ash percentage, whereas femurs were evaluated for bone mineral density (BMD), bone mineral content, bone volume as a fraction tissue volume, and porosity percentage from total, cortical, medullary, and trabecular bones. The higher TBS (P = 0.0005) and ash percentage (P = 0.045) was observed in hens raised in FR systems compared with those raised in the CC. Overall, TBS of W-36 hens was significantly greater than that of HB hens (P < 0.0001); however, there was no difference in the ash percentage between the strains (P > 0.05). An interaction between the housing environment and hen strain was observed for BMD (P = 0.04), wherein W-36 hens raised in the FR system had higher BMD than HB hens. Similarly, hens raised in FR systems had higher trabecular bone volume than those raised in CC (P = 0.022). Hen strain influenced total and cortical bone properties: BMD, bone volume as a fraction tissue volume, and porosity percentage, wherein W-36 hens had better properties than HB hens (P < 0.05). Trabecular BMD was higher in W-36 hens than in HB hens (P = 0.04), whereas bone volume was higher in HB hens (P < 0.0001). The results suggest that raising laying hens in alternative housing systems that have provision for exercise such as FR reduces structural bone loss, stimulate structural bone formation, and improve breaking strength of bones; however, it varies with the strain.

Evaluation of Alternative Euthanasia Methods of Neonatal Chickens.

Hatched male layer chicks are currently euthanized by maceration in the United States. Public concerns on the use of maceration have led to the search for alternative methods. We hypothesized that gas inhalation and low atmospheric pressure stunning (LAPS) are viable and humane alternatives to instantaneous mechanical destruction. The objective of this study was to evaluate the physiological and behavioral responses of recently hatched male layer chicks when subjected to carbon dioxide, nitrogen inhalation, or LAPS. The study consisted of seven treatments: breathing air (NEG), 25% carbon dioxide (CO2), 50% CO2, 75% CO2, 90% CO2, 100% nitrogen (N2), or LAPS. Ten day-of-hatch, male layer chicks were randomly assigned to each treatment, and each treatment was replicated on ten different days. A custom-made vacuum system was used to reduce air pressure inside the chamber from 100.12 kPa to 15.3 kPa for the LAPS treatment. Serum corticosterone and serotonin levels were measured using commercially available competitive enzyme linked immunosorbent assay (ELISA). Latencies to loss of posture and motionlessness were determined from video recordings. The 25% and 50% CO2 treatments were discontinued after the first replication, as the majority of the chicks recovered. The chicks in the negative (NEG) group had significantly higher levels of corticosterone than the other four euthanasia treatments. On the other hand, the serotonin levels of chicks in the NEG group was significantly lower when compared to the other four euthanasia treatments. The latencies to loss of posture and motionlessness of chicks exposed to 75% and 90% CO2 were significantly shorter than those in the LAPS and N2 inhalation treatments. These data suggest that the stress responses of chicks to the CO2, N2, and LAPS treatments do not differ among each other. However, the CO2 inhalation method was faster in inducing loss of posture and motionlessness in chicks than the LAPS and N2 inhalation treatments.

Carbon Dioxide and Nitrogen Infused Compressed Air Foam for Depopulation of Caged Laying Hens.

Depopulation of infected poultry flocks is a key strategy to control and contain reportable diseases. Water-based foam, carbon dioxide inhalation, and ventilation shutdown are depopulation methods available to the poultry industry. Unfortunately, these methods have limited usage in caged layer hen operations. Personnel safety and welfare of birds are equally important factors to consider during emergency depopulation procedures. We have previously reported that compressed air foam (CAF) is an alternative method for depopulation of caged layer hens. We hypothesized that infusion of gases, such as carbon dioxide (CO2) and nitrogen (N2), into the CAF would reduce physiological stress and shorten time to cessation of movement. The study had six treatments, namely a negative control, CO2 inhalation, N2 inhalation, CAF with air (CAF Air), CAF with 50% CO2 (CAF CO2), and CAF with 100% N2 (CAF N2). Four spent hens were randomly assigned to one of these treatments on each of the eight replication days. A total of 192 spent hens were used in this study. Serum corticosterone and serotonin levels were measured and compared between treatments. Time to cessation of movement of spent hens was determined using accelerometers. The addition of CO2 in CAF significantly reduced the foam quality while the addition of N2 did not. The corticosterone and serotonin levels of spent hens subjected to foam (CAF, CAF CO2, CAF N2) and gas inhalation (CO2, N2) treatments did not differ significantly. The time to cessation of movement of spent hens in the CAF N2 treatment was significantly shorter than CAF and CAF CO2 treatments but longer than the gas inhalation treatments. These data suggest that the addition of N2 is advantageous in terms of shortening time to death and improved foam quality as compared to the CAF CO2 treatment.