The scientific community needs objective measures to appropriately assess animal welfare. The study objective was to assess the impact of housing system on novel physiological and behavioral measurements of animal welfare for laying hens, including secretory and plasma Immunoglobulin (IgA; immune function), feather corticosterone (chronic stress), and attention bias testing (ABT; anxiety), in addition to the well-validated tonic immobility test (TI; fearfulness). To test this, 184 Bovan brown hens were housed in 28 conventional cages (3 birds/cage) and 4 enriched pens (25 birds/pen). Feces, blood, and feathers were collected 4 times between week 22 and 43 to quantify secretory and plasma IgA and feather corticosterone concentrations. TI tests and ABT were performed once. Hens that were from cages tended to show longer TI, had increased feather corticosterone, and decreased secretory IgA at 22 weeks of age. The caged hens fed quicker, and more hens fed during the ABT compared to the penned hens. Hens that were in conventional cages showed somewhat poorer welfare outcomes than the hens in enriched pens, as indicated by increased chronic stress, decreased immune function at 22 weeks of age but no other ages, somewhat increased fear, but reduced anxiety. Overall, these novel markers show some appropriate contrast between housing treatments and may be useful in an animal welfare assessment context for laying hens. More research is needed to confirm these findings.

Many benefits have been found in supplementing essential oils such as oregano oil (EOO) to poultry, including increased body weight gain, antioxidant activity, and better gastrointestinal morphology. However, few studies tested the influence of EOO supplementation on laying hens and reported conflicting results regarding its efficacy in improving their health and performance. Therefore, we aimed to explore the effects of dietary EOO on performance, gastrointestinal (GIT) traits, blood lipid, and antioxidant capacity in laying hens during the rearing phase. A total of 300-day-old Hy-line-Brown chicks were used, and treatment diets consisted of corn-soybean based either without (CON) or with EOO (Ecodiar®, 0.275 g/kg diet). Birds were randomized across treatments with five pens/treatment and 30-birds/pen. Pen weights and feed rejected were recorded every two weeks (1-17 weeks of age), to calculate daily feed intake (ADFI), body weight (BW), and daily weight gain (ADWG). At 11 and 14 weeks of age, blood samples were collected from 3 birds/pen and analyzed for blood lipids and antioxidant levels, and 5-birds/treatment were euthanized, and GIT traits were tested. Differences in measured parameters across weeks and between treatments were assessed using GLMM with Tukey’s Post hoc test applied to significant results in R 3.3.1 (α set at 0.05). Body weights at weeks 3, 11, 13, and 17 were significantly higher in the EOO group compared to the CON group (all P ≤ 0.05), ADWG was significantly higher in EOO birds compared to CON birds at 9 and 13 weeks old (all P ≤ 0.05), while no significant differences in ADFI were observed between treatments across weeks of the trials. At both 11 and 17 weeks old, triglyceride levels were significantly lower, while high-density-lipoprotein levels were higher in EOO (all P ≤ 0.05). Malondialdehyde levels were lower in the EOO group versus CON (p=0.01), while EOO birds had higher glutathione levels (p=0.01) than CON. Finally, at 12 weeks old, the weight of the entire GIT and empty gizzard were higher in the EOO group versus CON (all P ≤ 0.05), while liver and spleen weights were not significantly different between groups. In conclusion, dietary oregano supplementation exerted promoting effects on the performance of Hy-Line Brown pullets.

A complex system of neural pathways, collectively known as the microbiota–gut–brain (MGB) axis, interconnects the gut microbiota, the gastrointestinal system, and the brain along with its periphery. Previous studies have demonstrated that modulation of the MGB axis can influence stress-related behaviors such as anxiety. This connection becomes apparent in scenarios like agonistic behavior in laying hens, which is characterized by aggressive head and feather pecks, that can ultimately result in cannibalism and death. The objective was to examine the effects of a dietary synbiotic on agonistic behavior, plasma and brain monoamines, stress parameters, and cecal microbiota counts via modulation of the MGB axis. A total of 396 W36 Hy-Line laying hens were provided at random with a control (CON: basal diet) or treatment (SYN: basal diet supplemented with synbiotic) diet from 50 to 60 weeks old (nine pens/treatment, 22 birds/pen). Blood samples and video recordings (three consecutive days/week) were taken at 50 and 60 weeks. At 60 weeks, three hens/pen were euthanized for brain and cecal microbiota collection. Threatening, fighting, head, body, and feather pecking all occurred less frequently at 60 weeks in the SYN group (p < 0.05). Plasma corticosterone, adrenocorticotropic hormone, dopamine, and serotonin were significantly lower while tryptophan and 5-hydroxyindoleacetic acid were significantly higher in birds from the SYN group (p < 0.05). Significant differences in serotonin, 5-hydroxyindoleacetic acid, dopamine, homovanillic acid, and 3,4-dihydroxyphenylacetic acid were observed in the hypothalamus, hippocampus, and amygdala of the brain. Serotonin and dopamine turnover rates were significantly different in all three regions of the brain (p < 0.05). Cecal counts of Lactobacillus and Bifidobacterium were significantly higher in the SYN group (p < 0.05). Synbiotic supplementation resulted in many significant differences, indicating activation of the serotonergic systems and modulation of both the MGB axis and HPA axis with positive effects on welfare and stress.

Prior research suggests exercise during pullet rearing can mitigate lay-phase bone fractures by strengthening muscles, enhancing balance, and increasing bone mass. This study aimed to confirm that Hy-Line brown pullets with multi-tier perches show increased activity and improved musculoskeletal health. Pullets (n = 810) were randomly allocated to housing systems, either with multi-tier perches (P; n = 15 pens) or without (NP; n = 15 pens), spanning from 0–17 weeks of age. At 5, 11, and 17 weeks, individual birds were meticulously monitored for activity using accelerometers over three consecutive days (n = 90 randomly selected birds/week). At 11 and 17 weeks, 60 birds underwent euthanasia and computed tomography (CT) scans to ascertain tibiotarsal bone mineral density and cross-sectional area measurements. Post-CT scanning, birds were dissected for muscle size, tibiotarsal breaking strength, and tibiotarsal ash percentage measurements. Additionally, serum concentrations of bone-specific alkaline phosphatase and procollagen type 1 N-terminal propeptide were assessed as markers of bone formation (n = 90 birds/week). Pullet group P exhibited heightened vertical activity (P0.05) during weeks 5, 11, and 17 compared to group NP. Tibiotarsal bones of P pullets demonstrated superior total and cortical bone mineral density at week 11, alongside increased cortical bone cross-sectional areas and heightened total and cortical bone mineral densities at week 17 (P<0.05) compared to NP pullets. At week 11, P pullets displayed larger leg muscles, including triceps, pectoralis major and minor, and leg muscles at week 17 (P<0.05) compared to NP pullets. Notably, at both weeks, P pullets’ tibiae exhibited greater breaking strengths, higher ash percentages, and elevated concentrations of bone-specific alkaline phosphatase and procollagen type 1 N-terminal propeptide compared to NP pullets (P<0.05). The study findings underscore the benefits of providing multi-tier perches for pullets, serving as a valuable tool for enhancing bird activity and musculoskeletal health preceding the lay phase.

Boron supplementation may improve the musculoskeletal health of pullets before entering the lay phase. This study aimed to evaluate the effects of different boron amounts on the performance, muscle deposition, tibia cross-sectional area (CSA) and mineral density (BMD), ash percent, breaking strength, and bone mineralization (bone-specific alkaline phosphatase [BALP] and pro-collagen type 1 n-terminal propeptide [P1NP]) of a white-feathered strain of pullets. A total of 528 Hy-Line W-36 pullets were distributed across 24 pens and fed basal diets containing varying amounts of boron (C: 0 mg/kg; L: 50 mg/kg; M: 100 mg/kg; H: 150 mg/kg) for 17 weeks. Performance parameters (body weight, average daily weight gain/bird, and average daily feed intake/bird) were measured at weeks 4, 7, 10, 13, and 16, while all other measures were taken at 11 and 17 weeks of age. Performance was not impacted by boron supplementation. Pectoralis major weights were higher in H pullets at 11 weeks of age, and we also observed higher pectoralis major, minor, and leg muscle weights in H pullets at 17 weeks of age. Pullets fed the H diet had larger cortical CSA than the other treatment groups at 11 weeks of age. At 17 weeks of age, both the H and M groups had larger cortical CSA than the L and C groups, but the M group had slightly smaller cortical CSA. Pullets fed the H diet had higher BMD values than the other treatment groups at 11 weeks of age. At 17 weeks of age, pullets fed the H diet had the highest total BMD values compared to the other treatment groups, and cortical BMD increased with increasing boron inclusion. Pullets fed the H diet had the highest tibia ash percentages and concentrations of BALP and P1NP. Pullets fed the M and H diets had greater failure load and maximum bending moment than pullets fed the L or C diet at 11 weeks of age, with H pullets having greater stiffness values than other groups. At 17 weeks of age, pullets fed the H diet had greater failure load and maximum bending moment compared to all other treatment groups. Our results suggest that providing boron within the diet at 150 mg/kg can improve the musculoskeletal characteristics of Hy-Line W-36 pullets up to 17 weeks of age, without impacting performance parameters.

Perches can enhance laying hen welfare, but their effectiveness might be age-dependent. We investigated early and late perch access effects on anxiety and fear in pullets through attention bias (AB) and tonic immobility (TI) tests. Pullets (n = 728) were raised with or without multi-level perches: CP (continuous perch access: 0–37 weeks), EP (early perch access: 0–17 weeks), LP (late perch access: 17–37 weeks), and NP (no perch access). AB was conducted in weeks 21 and 37 (n = 84/week), and TI was performed in weeks 20, 25, and 37 (n = 112/week). CP hens fed quicker than EP, LP, and NP in AB at weeks 21 and 37 (p ≤ 0.05). CP and NP feeding latencies were stable, while EP and LP fed faster at week 37 (p ≤ 0.05). CP had the shortest TI at week 20 (p < 0.05). CP and LP had the shortest TI in weeks 25 and 37 (all p ≤ 0.05). Unlike NP, CP reduced anxiety and fear. Adding perches during laying (LP) raised anxiety at week 21, adapting by week 37, and removing pre-laying perches (EP) worsened fear at weeks 20 and 25 and anxiety at week 21, recovering by week 37. Adding or removing perches prior to the lay phase increased fear and anxiety, an effect that disappeared by week 37 of age. Our study indicates that continuous perch access benefits animal welfare compared to no perch access at all.