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Adipokines including adiponectin (ADIPO), chemerin (CHEM) and visfatin (VISF) are involved in metabolism and reproductive functions. These 3 adipokines are present in ovarian cells in different preovulatory follicles in hens. We have previously shown that VISF and ADIPO are able to modulate in vitro steroid production by hen granulosa cells (GCs). It is, however, unclear whether CHEM acts on hen ovarian cells. In addition, no study has yet investigated the effect of a mixture of several adipokines such ADIPO, VISF, and CHEM on GCs from different preovulatory follicles. In this study, we investigated the effect of CHEM alone and in combination with ADIPO and VISF on cell viability, proliferation and progesterone secretion in cultured granulosa cells (GCs) from the largest follicles F1 and smaller ones (F3/F4) in the presence of gonadotropins (oLH and oFSH) or hIGF-1. First, various concentrations of chemerin were examined (0, 12, 25, 50, and 100 ng/mL) and then we determined the response to CHEM (at 25 ng/mL) in combination with ADIPO (10 µg/mL) and VISF (100 ng/mL). Chemerin exposure did not affect F1 and F3/F4 granulosa cell viability and proliferation whatever the concentation and in the presence of the mixture. However, it reduced progesterone secretion in dose dependent manner in both F1 and F3/F4 follicles. Furthermore, this CHEM inhibitory effect was significantly higher when CHEM was combined with ADIPO and VISF. Furthermore, CHEM reduced significantly oLH and oFSH- induced progesterone secretion in F1 GCs and oFSH and hIGF-1-induced progesterone secretion in F3/F4 GCs. Interestingly, this inhibitory effect of CHEM was similar in F1 GCs when CHEM was in mixture with ADIPO and VISF whereas it was significantly higher in F3/F4 GCs. Taken together, CHEM impairs progesterone secretion in cultured hen GCs and this inhibitory effect can be potentiated when it is in combination with other adipokines., Competing Interests: DISCLOSURES The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Avian mycoplasmas are known pathogens, which cause severe economic losses in poultry flocks. PCR is a rapid, sensitive, and less expensive diagnostic tool than culture for the identification of mycoplasmas in poultry farms. The objective of this study was to determine by PCR the presence of Mycoplasma spp., Mycoplasma gallisepticum (MG), Mycoplasma synoviae (MS), and Mycoplasma pullorum (MP) in laying hens located in the Sfax region, in the South of Tunisia., Results: A total of 781 tracheal swabs were collected from 13 laying-hen farms without clinical signs at the date of sampling. MP was detected by a newly described specific PCR assay. The prevalence calculated from PCR results at the flock level was 100% for Mycoplasma spp., 0% for MG, 84.6% for MS and 61.5% for MP. The overall prevalence at the animal level was 38.7% for Mycoplasma spp., 0% for MG, 25% for MS and 6.4% for MP. The overall prevalence of 100% of avian mycoplasmas in laying-hen farms (38.7% prevalence at the animal level) shows an alarming situation., Conclusions: These results underline the importance of monitoring the emergence and spread of Mycoplasma strains in farms in order to decrease economic losses due to mycoplasmoses., (© 2024. The Author(s).)

The aim of this study was to determine the effect of oligosaccharide extract from bamboo shoot (BOS) on the performance, intestinal morphology, microbiota and immune reaction of laying hens challenged with dextran sodium sulfate (DSS). Thirty-two White Leghorn hens (480 days old) were divided into 4 groups (8 hens each) with similar mean body weights: C (basal diet), D (basal diet + DSS), B (5 g/kg BOS diet), and BD (5 g/kg BOS diet + DSS). They were administered a single oral dose of 4 mL of distilled water/kg body weight with or without 0.45 g of DSS for 7 consecutive days from the 14th d to the 21st d of the experiment. The important findings were that (1) The egg yolk ratio was decreased by DSS treatment, but it was improved by BOS treatment, which also increased the egg shell ratio. (2) The diversity of intestinal microbiota and relative abundance of 4 bacteria genera were increased by BOS treatment. (3) Intestinal morphology was not affected by DSS and BOS, but the leukocytes accumulation in the liver was increased by DSS treatment and suppressed by BOS treatment. (4) Dietary BOS treatment influenced the mRNA expression of Th-1 and Treg cytokines in the liver and Th-17 cytokines in both intestine and liver of laying hens. These results suggest that BOS may enhance egg quality, Th-1 and Th-17 immune function without causing tissue damage under normal condition, and may suppress the excessive inflammatory responses during inflammation., Competing Interests: DISCLOSURES The authors have no conflicts of interest directly relevant to the content of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The skeletal health of laying hens improves when birds are given opportunities to perform load-bearing movements with elevated structures, such as perches. We investigated how early access to elevated structures varying in complexity and height would affect bone quality and subsequent keel bone fractures in a layer multitiered aviary. Female Dekalb White pullets were reared in floor pens furnished with floor perches (FL), single-tiered aviaries (ST), or 2-tiered aviaries (TT; n = 5 pens/treatment) through 16 wk of age. At 17 wks, all structures were replaced with identical multitiered layer aviaries. The keel, both tibiae, and both humeri were collected from 60 euthanized birds from each rearing treatment at 8, 16 and 30 wk of age, and analyzed with dual X-ray absorptiometry (DEXA) for bone mineral density and length. At 18, 26, 28, and 30 wk of age, 10 focal hens/pen were radiographed repeatedly and the presence, severity of keel bone fractures were assessed with a tagged visual analogue scale. The number of fractures was also recorded. At 16 wk of age, FL pullets had lower BMD of the tibia (P = 0.003), keel (P = 0.013), and humerus (P = 0.004) compared to ST and TT pullets. Most of the observed treatment differences disappeared after pullets were transferred to the aviary. BMD continued to increase for all hens through 30 wk of age. Pullet rearing did not affect the presence or severity of keel bone fractures, or number of new fractures incurred between ages (P > 0.05). The prevalence and severity of keel bone fractures increased between 26 to 28 wk and remained high to 30 wk of age (P < 0.0001). Hens experienced more new fractures between 26 to 30 wk than between 18 to 26 wk of age (P = 0.0046). The effects of pullet housing on bone quality were short-term when hens had access to adult housing with multiple opportunities for load-bearing movements. Keel fractures with minor severity were high in prevalence reflecting the use of radiography to assess this injury., Competing Interests: DISCLOSURES The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Maja Makagon reports financial support was provided by Foundation for Food and Agriculture Research. Maja Makagon reports equipment, drugs, or supplies was provided by Lubing. Richard Blatchford reports was provided by Foundation for Food and Agriculture Research. Richard Blatchford reports equipment, drugs, or supplies was provided by Lubing. John Tarlton reports financial support was provided by Foundation for Food and Agriculture Research. Michael Toscano is a Section Editor for Poultry Science; Maja Makagon is an Associate Editor for Poultry Science If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The effect of the diet modification (soybean and lupine addition) on the content of protein and amino acids (AA) in eggs was studied. Both the sampling day and the diet influenced the total protein content. In albumen, the lowest protein content (10.6%) was noted after administering a diet containing 25% lupine; in the same egg the yolk contained the most proteins (16.7%). In the content of nonessential AA (NAA) in egg yolks, differences were noted only for cysteine, with its the highest content in the yolks of the control group. The stable content of essential yolk amino acids (EAA) was observed only for isoleucine, leucine, tryptophan and phenylalanine. The highest contents of EAA and NAA were recorded in the yolks of the control group (~47 and ~53 g/100 g of protein, respectively) and in the group with 25% additions of lupine (~42 and ~51 g/100 g of protein, respectively). AA with constant content in the tested albumens were methionine, tryptophan and alanine. The highest content of EAA (>~42 g/100 g of protein) and NAA (>~62 g/100 g of protein) were determined in albumen of eggs determined in the group with at least 20% additions of lupine. The highest content of EAA for humans delivered eggs from groups 4-6 (with the addition of soy into the diet ≤5%). The protein sources used in the hen diet significantly influenced the content of protein and individual AA in the produced eggs.

Despite the plethora of studies on the impacts of access to runs on chickens, there is a paucity of information on the welfare and behavioural repertoire of hens raised in the deep litter houses with or without access to legume- or grass-based pasture. Therefore, this study aimed to evaluate the impact of access to grass or legume pastures by laying hens on behaviour, physiological responses and bacterial load. The study was conducted to evaluate the influence of exposure of egg-type chickens to runs on grass or legume pastures on their welfare and behaviours. The study involved the use of 240 ISA brown pullets from 12 weeks of age and and lasted for 48 weeks. The treatments were deep litter housing with grass-based pasture run (PG), deep litter housing with legume-based pasture run (PL) and deep litter housing without runs (LD) having 80 pullets with four replicates of twenty birds each. Behavioural observations of the hens in each pen were made at 52 weeks of age and tonic immobility was assessed by making the birds lie on their back with their head resting in a U-shaped wooden cradle. The measurements of the respiratory rate and rectal temperature of the hens were assessed at 1:00 p.m. at different laying phases. The gastrointestinal and egg bacterial counts were conducted at 60 weeks of age. Results revealed that the proportion of time spent eating was highest (p < 0.05) in the deep litter housing system, while the legume and grass pasture were similar. The hens spent most of their time standing and eating in the three treatments. However, the time spent standing in PL and PG was similar but significantly higher (p < 0.05) than in LD. Results on tonic immobility duration showed that the time spent by the hens in LD during the reaction was significantly longer than those of the PL and PG in the first, second and third phases of the experiments. However, the time spent by the hens in PL and PG was similar. The rectal temperatures of PL and PG birds were comparable and higher than those of LD during the second phase. On the other hand, there was no difference in the respiratory rate. Plasma triiodothyronine (T 3 ) of the hens did not follow a consistent pattern. The bacterial count in the large intestine in LD and PL was similar but significantly (P < 0.05) higher than that of the PG. It was concluded that access to pasture influenced the behaviours of hens and that tonic immobility duration was shorter in the hens on the pasture, suggesting that access to pasture favoured hens' welfare., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Background: The benefits of home enteral nutrition (HEN) are increasingly recognized, with more scholars focusing on this field. This study aimed to comprehensively identify collaborative networks, analyze, and track research trends, focus on current hotspots, and accurately predict the forefront and focus of home enteral nutrition., Methods: A computer search of the Web of Science Core Collection (WoSCC) was conducted for studies related to home enteral nutrition published from January 1, 2004, to December 31, 2023, and select them in compliance with the PRISMA guidelines. The CiteSpace software was used for bibliometric visualization and comparative analysis of countries, institutions, journals, references, and keywords., Results: A total of 1,113 documents were included, showing a steady annual increase in publication volume. The United States and the Mayo Clinic were the top publishing country and institution, with 302 and 41 papers, respectively. "CLIN NUTR" had the highest number of publications, totaling 221, while "ESPEN guideline on home enteral nutrition" was the most cited reference, with 43 citations. The most prolific author was Manpreet S with 29 papers., Conclusion: The management of HEN is a current research hotspot. The safety of HEN and how to improve patient compliance are critical areas for researchers to consider. Future research could focus on these aspects. The blurring of boundaries between hospital and home care and how to utilize telemedicine technologies to serve more patients deserve in-depth exploration. Researchers worldwide should combine their unique characteristics and advantages to strengthen international cooperation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Qin, Chen, Huang, Xu, Hu, Suo, Hu and Peng.)

Background: Given the growing use of home enteral nutrition (HEN), assessing the experience of consumers and caregivers is crucial to understanding the real-world subjective and objective challenges of administering HEN., Methods: After obtaining institutional review board approval, a survey was distributed to HEN consumers and caregivers between January 16, 2020, and July 16, 2021. Data collected included information regarding demographics, primary diagnosis, tube and connectors, HEN regimen, and overall HEN experience., Results: A total of 884 individuals responded to the survey: 673 (76.1%) responses by caregivers and 211 (23.9%) responses by patients. The study cohort included 566 (64%) children and 318 (36%) adults. The leading primary diagnosis of participants was developmental delay and motility disorder for children and adults, respectively. Low-profile gastric tubes were the most used (75.7% of children and 30.3% of adults). Notably, legacy connectors were utilized for more patients (46.7% children, 52.6% adults) compared to ISO-80369-3 connectors (38.9% children, 29.7% adults). HEN complications were prevalent, including enteral tube site infections and other tube-related complications, including clogging and kinking., Conclusion: This real-world data reveals that HEN complications remain prevalent. Additionally, despite introducing ISO-80369-3 connectors many years ago, most patients continue to use legacy tubes with a significant lack of knowledge about ISO-80369-3 connectors. The survey results guide HEN providers to focus on several areas to reduce complications., Competing Interests: Conflicts of interest Authors declare no relevant conflict of interest., (Copyright © 2024 European Society for Clinical Nutrition and Metabolism. Published by Elsevier Ltd. All rights reserved.)

The decrease in eggshell quality seriously affects production efficiency. Guinea fowl (GF) eggs possess strong eggshells because of their unique crystal structure, and few systematic studies have compared laying hen and GF eggs. Sixty eggs were collected from both 40-week-old Dwarf Layer-White (DWL-White) laying hens and GF, and the eggshell quality, ultrastructure, bubble pores, and composition were measured. The results showed that the DWL-White eggs had a higher egg weight and a lower eggshell strength, strength per unit weight, thickness, and ratio than the GF eggs ( p < 0.01). There were differences in the mammillary layer thickness ratio, the effective layer thickness ratio, the quantity of bubble pores (QBPs), the ratio of the sum of the area of bubble pores to the area of the eggshell in each image (ARBE), and the average area of bubble pores (AABPs) between the DWL-White and GF eggs ( p < 0.01). The composition analysis demonstrated that there were differences in the organic matter, inorganic matter, calcium, and phosphorus between the DWL-White and GF eggs ( p < 0.01). There were positive associations between the mammillary knob number in the image and the QBPs and ARBE and a negative correlation with the AABPs in the DWL-White eggs ( p < 0.01). This study observed distinctions that offer new insights into enhancing eggshell quality.

A random-effects meta-analysis was conducted to investigate the effect of mycotoxins (MT) without or with the inclusion of yeast cell wall extract (YCWE, Mycosorb ® , Alltech, Inc., Nicholasville, KY, USA) on laying hen performance. A total of 25 trials were collected from a literature search, and data were extracted from 8 of these that met inclusion criteria, for a total of 12 treatments and 1774 birds. Laying hens fed MT had lower ( p < 0.05) body weight (BW) by -50 g, egg production by -6.3 percentage points, and egg weight by -1.95 g than control fed hens (CTRL). Inclusion of YCWE during the mycotoxin challenges (YCWE + MT) resulted in numerically greater ( p = 0.441) BW by 12.5 g, while egg production and egg weight were significantly ( p < 0.0001) higher by 4.2 percentage points and 1.37 g, respectively. Furthermore, economic assessment calculations indicated that YCWE may not only support hen performance but also resulted in a positive return on investment. In conclusion, mycotoxins can play a role in negatively impacting laying hen performance and profitability. Inclusion of YCWE in feed with mycotoxin challenges provided benefits to egg production and egg weight and may support profitability. As such, the inclusion of YCWE could play an important role in minimizing mycotoxin effects and in turn aid farm efficiency and profitability.

Due to the intensive development of novel biopharming applications, there is a need for the in vitro verification models prior to in vivo testing. Laying hen has been already applied as an animal bioreactor to produce the therapeutical enzyme in a rare disease called lysosomal acid lipase deficiency. In this study, we aimed to verify how the proteome of the transfected oviduct epithelial cells would be affected by genetic nonviral modification with the human exogene. The study was based on a previously developed method to cultivate chicken oviduct epithelial cells (COEC). The typical characteristics of the COEC epithelial cells were retained across the experiments. The mean efficiency of nucleofection ranged from 2.6 to 19.7% depending on the cells' isolation and location in the oviduct (upper, infundibulum site, or magnum). The PCR confirmed the incorporation of human interferon alpha2a (hIFNα2a) exogene into the nucleofected COEC but, the production of hIFNα2a protein did not exceed the detection level in this study. The ovalbumin protein was detected in the nontransfected and transfected COEC, which confirmed the normal secreting functions of the cells subject to modification. Proteomic analysis revealed an increase in abundance of the cell adhesion molecules and collagen molecules after introducing gene under ovalbumin promoter. According to the bioinformatic analyses there was a limited negative impact of transfection on cells, and the normal biochemical pathways were not severely disordered. In conclusion, the observations provide new knowledge about the proteomic profile of the manipulated COEC with regard to the retained normal functionality of the cells, which can be informative for avian biopharma research., (Copyright © 2024. Published by Elsevier Inc.)

Infectious bronchitis virus (IBV) is a respiratory virus causing atropism in multiple body systems of chickens. Recently, the California 1737/04 (CA1737/04) IBV strain was identified as one of the circulating IBV variants among poultry operations in North America. Here, the pathogenicity and tissue tropism of CA1737/04 IBV strain in specific-pathogen-free (SPF) hens were characterized in comparison to Massachusetts (Mass) IBV. In 30 weeks-old SPF hens, Mass or CA1737/04 IBV infections were carried out, while the third group was maintained as a control group. Following infection, we evaluated clinical signs, egg production, viral shedding, serology, necropsy examination, and histopathology during a period of 19 days. Also, certain tissue affinity parameters were investigated, which involved the localization of viral antigens and the detection of viral RNA copies in designated tissues. Our findings indicate that infection with CA1737/04 or Mass IBV strain could induce significant clinical signs, reduced egg production, and anti-IBV antibodies locally in oviduct wash and systemically in serum. Both IBV strains showed detectable levels of viral RNA copies and induced pathology in respiratory, renal, enteric, and reproductive tissues. However, the CA1737/04 IBV strain had higher pathogenicity, higher tissue tropism, and higher replication in the kidney, large intestine, and different segments of the oviduct compared to the Mass IBV strain. Both IBV strains shed viral genome from the cloacal route, however, the Mass IBV infected hens shed higher IBV genome loads via the oropharyngeal route compared to CA1737/04 IBV-infected hens. Overall, the current findings could contribute to a better understanding of CA1737/04 IBV pathogenicity in laying hens., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Ali, Farooq, Altakrouni, Najimudeen, Hassan, Isham, Shalaby, Gallardo and Abdul-Careem.)

The objective of this study was to identify and compare the quality characteristics and concentrations of various compounds in eggs from several pure breeds and lines of hens reared under the same environmental conditions and fed a commercial feed. A total of 280 hens aged 52 to 56 wk belonging to 14 different breeds or lines of hens worldwide were included in this study. Their eggs were characterized by wide differences in various egg quality parameters. Breeds and lines of hens with a higher lutein content in eggs were characterized by a lower beta-carotene content (e.g. Hy line brown, Cochin miniature, Ayam Cemani) (P < 0.001). Additionally, vitamin D, cholesterol, and fatty acid contents were also different between eggs, from 1.51 to 1.79 μg/100g; from 14.1 to 15.4 mg/g fat, PUFA from 19.6 to 22.8 g/100g fat, and SFA from 32.8 to 37.8 g/100g fat respectively (P < 0.001). Lysozyme content also exhibited significant variation among breeds, with some showing a 2-fold higher content in eggs compared to others (0.31% - cochin miniature, 0.66% Faverolle) (P < 0.001). Our study demonstrated that intensively selected hen breeds like Hy-line Brown Hybrid had an improved egg quality seen by the increase in many parameters (e.g., egg weight, Haugh unit, Lutein, vitamins D, MUFA) compared to pure breed hens. In conclusion, genetic differences between breeds and lines of hens have a significant impact on the quality of eggs., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Follicle selection in hens refers to a biological process that only one small yellow follicle (SYF) is selected daily or near-daily for following hierarchical development (from F5/F6 to F1) until ovulation. MFN2 is a kind of GTPases located on the mitochondrial outer membrane, which plays a crucial role in mitochondrial fusion. This study aimed to elucidate the role of MFN2 in proliferation and progesterone biosynthesis of granulosa cells (GCs) during follicle selection in hens. The results showed that GCs began to produce progesterone (P4) after follicle selection, accompanied with changes from multi-layer with flat cells to single layer with cubic cells. MFN2 was detected in GCs of follicles from SYF to F1. After follicle selection, the expression level of MFN2 in GCs upregulated significantly, accompanied with increases in P4 biosynthesis, ATP production, mitochondrial DNA (mtDNA) copy numbers of granulosa cells. FSH (80 ng/mL) facilitated the effects of P4 biosynthesis and secretion, ATP production, mtDNA copy numbers, cell proliferation and the MFN2 transcription of granulosa cells from F5 (F5G) in vitro. However, FSH treatment did not promote P4 secretion in granulosa cells from SYF (SYFG) in vitro. Meanwhile, we observed that change fold of MFN2 transcription, ATP production, mtDNA copy numbers and cell proliferation rate in F5G after treatment with FSH were greater than those in SYFG. Furthermore, expression levels of MFN2 protein and messenger RNA in F5G were significantly higher than those in SYFG after treatment with FSH. P4 biosynthesis, ATP production, mtDNA copy numbers as well as cell proliferation reduced significantly in F5G with MFN2 knockdown. Oppositely, P4 biosynthesis, ATP production, mtDNA copy numbers and cell proliferation increased significantly in SYFG after the overexpression of MFN2. Our results suggest that the upregulation of MFN2 may be involved in the initiation of P4 biosynthesis, and promotion of GCs proliferation during follicle selection., (© 2023 Wiley Periodicals LLC.)

When using chickens in animal studies, the handling of these animals for sample collection or general examinations is considered stressful due to their prey nature. For the study presented here, plasma and salivary corticosterone as well as New Area Test behavior and fecal output were used to evaluate whether it is possible to influence this stress perception using a three-week clicker training program. The results indicate that clicker training seems to be a suitable refinement measure in the sense of cognitive enrichment for the husbandry of this species. However, since it was also shown that three-week training was not sufficient to sustainably reduce the stress perception with regard to prolonged stressor exposure, and since it was also evident that manipulations such as routine blood sampling are perceived as less stressful than assumed, further studies with prolonged training intervals and situations with higher stressor potential are warranted. Also, further parameters for training assessment must be considered. For the general use of training as a supportive measure in animal experiments, its proportionality must be considered, particularly considering the expected stress and adequate training time.

Background: In February 2022, the US Food and Drug Administration published a safety communication regarding the risk of strangulation from enteral feeding delivery sets. When it comes to the causes of accidental strangulation or asphyxiation, it is common knowledge that household items, such as window blind cords, are potential hazards. However, many people may not expect that medical devices may pose similar risks because of medical line entanglement (MLE)., Methods: A survey of clinicians from pediatric acute care and outpatient facilities and caregivers of patients with medical lines was conducted with a goal to understand if clinicians and caregivers are aware of MLE, if healthcare facilities have policies and procedures related to the prevention of MLE, and if education about the risks of MLE is provided to caregivers when they first receive a medical device., Results: The survey was shared through clinician organizations and patient advocacy groups. Responses were received from 191 clinicians and 117 caregivers. A majority of clinicians reported awareness of the risk of entanglement; however, few received direction from their employer on managing these risks. Caregivers (N = 106) reported that their child had experienced MLE, however only 9% of those caregivers recalled receiving any education on MLE from their healthcare providers., Conclusion: This survey demonstrates the need for healthcare facilities to develop programs for managing risks of MLE and for healthcare teams and caregivers to discuss prevention methods whenever an at-risk patient is sent home with a medical device that poses a risk for entanglement., (© 2023 American Society for Parenteral and Enteral Nutrition.)

Background: Alterations of the uterine microbiome are closely associated with various intrauterine diseases and physiological conditions, which are well-established in mammals. However, as representative oviparous animals, the research on the uterine microbial ecosystem and its functions with physiological homeostasis is limited in chickens. Additionally, continuous egg-laying disrupts the oviducal immune defenses of aged hens, susceptible to pathogen invasion, causing poor egg quality and food-borne infections in humans. Here, we investigated aging-related changes in the oviduct microbial colonization and transmission from the gut to eggs and their roles in a hen model., Results: The results of 16S rDNA sequencing showed significant differences in the oviduct microbial composition between young (38 weeks) and aged (77 weeks) laying hens. SourceTracker analysis further revealed differences in the effects of microbial transmission on the oviducal microbiota between young and aged hens. Enhanced barrier defense with cell apoptosis suppression and cell cycle arrest of the uterus were observed in aged hens reducing microbial transmission from the lower to upper reproductive tract. In addition, a total of 361 significantly differential metabolites were identified using metabolomics in the aged uterine microbiota, especially in products of amino acid metabolism and biosynthesis of various secondary metabolites, which might have essential effects on cell apoptosis by regulating immune responses and cell cycle. Notably, antibiotics disrupted uterine microbiota by dietary intervention and direct perfusion did not retard aging-related physiological changes but further aggravated aging processes by disrupting the cell cycle and apoptosis., Conclusions: The microbiota continuum along the reproductive tract in aged birds differs from that in young birds, especially with a significant shift in the uterus. The aged uterine microbiota probably contributes to the regulation of cell cycle and apoptosis by microbial metabolites primarily involved in amino acid metabolism and biosynthesis of various secondary metabolites. These findings provide new insights into the roles of the reproductive tract microbiota in regulating the cell programming of the aged host, contributing to the exploration of the microbiome as a target for diagnosing aging health status and therapy for gynecological diseases in women. Video Abstract., (© 2023. The Author(s).)

Fatty liver hemorrhagic syndrome (FLHS) is the most common metabolic disease in laying hens. Morus alba L. (mulberry) leaf has the effect of regulating lipid metabolism. We evaluated the effects of dietary 3% mulberry leaf (MUL) supplementation in production performance, egg quality, and liver lipid deposition in laying hens. Differentially expressed genes and circRNAs in the liver were identified using whole-transcriptome sequencing. We also evaluated the effects of the MUL extract using an in vitro model of primary hepatocytes induced by free fatty acids and explored the role of key circRNAs in this process. Dietary supplementation with 3% MUL alleviated liver steatosis in laying hens, as shown by decreased fatty liver color score, relative liver weight (P < 0.01), and triglyceride levels (P < 0.05), and showed a tendency to reduce the mortality rate of laying hens (P = 0.09). In addition, mulberry leaf supplementation significantly reduced cholesterol content in egg yolk (P < 0.01). Dietary mulberry leaf supplementation downregulated the expression of genes involved in fatty acid and cholesterol biosynthesis, and upregulated the expression of fatty acid oxidation-related genes in the liver. CircACACA, which is derived from exons 2 and 3 of the acetyl-CoA carboxylase alpha (ACACA) pre-mRNA, was significantly reduced in the MUL group (P < 0.01). Upregulation of circACACA expression reversed the lipid-lowering effect of mulberry leaf extract by upregulating sterol regulatory element-binding proteins 1 c (SREBP-1c) and fatty acid synthase (FASN) (P < 0.05). Overall, mulberry leaf is an effective therapeutic strategy for FLHS in hens and can improve liver lipid metabolism by downregulating circACACA., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Enriching eggs with omega-3 fatty acids (n-3 FA), such as docosahexaenoic acid (DHA), is a well-accepted practice that benefits the egg industry and consumers. However, issues around cost, sustainability, and product acceptance have necessitated the search for alternatives to feeding hens fish oil for DHA enrichment. The effects of feeding 2 algal oils on egg production and DHA enrichment in eggs and selected tissues were investigated. The algal oils were: 1) OmegaPro (OPAO) standardized algal oil for DHA content and 2) Crude algal oil (CAO). A total of 400, 46-wk-old Lohmann LSL lite hens were housed in enriched cages (10 birds/cage) and allocated 5 diets (n = 8) for a 12-wk trial. The iso-caloric and -nitrogenous diets were a standard corn and soybean meal diet, standard plus 0.25 or 0.76% OPAO and standard plus 0.23 or 0.69% CAO; algal oils diets supplied similar DHA at each level. Egg production indices (hen day egg production, feed intake, FCR, egg weight, egg mass, and eggshell quality) were monitored for 10 wk. Diet samples were analyzed for fatty acids (FA) on wk 1, 6, and 12 and eggs on wk 4, 5, 6, 9, and 12. At the end of the trial, one hen/cage was weighed and dissected for liver, breast and thigh for FA and long bones for ash content analyses. Concentration of omega-6 to omega-3 FA ratio was 12.9, 6.64, 3.48, 6.96, and 3.59 for standard, 0.23 and 0.76% OPAO, 0.25 and 0.69% CAO, respectively. Algal oils increased (P ≤ 0.046) eggshell thickness linearly. The concentration of DHA in the eggs from the birds fed the standard, 0.23 and 0.76% OPAO, 0.25 and 0.69% CAO was 84, 195, 286, 183, and 297 mg/100g egg, respectively, and algal oils enriched eggs with DHA linearly and quadratically (P ≤ 0.01). In conclusion, algal oils increased the concentration of DHA in eggs and had no adverse effects on egg production and eggshell quality., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

The month of hatching and the rearing management, especially temperature and photoperiod, are important factors for pullets and hens reared outdoors. The yield performance and egg quality of dual-purpose chicken breeds from the Veneto region (Italy), Pepoi (PP), Ermellinata di Rovigo (ER), Robusta Maculata (RM) and Robusta Lionata (RL), with different adult body weights (ABW, kg, PP = 1.3; ER = 2.3, RM and RL = 3.1), were studied, using a factorial model (4 × 2), considering breed and age (26-33 weeks, first age, summer-autumn, under decreasing natural photoperiod-on average, 12L:12D, and 42-53 weeks, second age, winter, under implemented photoperiod-14L:10D) as the main effects and interaction. The chicks hatched in spring, and they started laying at the end of summer/beginning of autumn. Significant ( p < 0.05) results were shown for many traits. ER showed higher hen-day egg production than that of PP, and RM and RL were the lowest; ER, RM and RL showed medium-size eggs and PP showed small-size eggs. RM produced the most spherical eggs and ER the most ovoid, and they showed the highest and the lowest eggshell thickness, respectively. RM showed the highest yolk to albumen ratio, and RL showed the lowest. The age increased the laying rate and the egg weight in all the groups. At 26-33 weeks, ER and PP showed higher hen-day egg production (on average 24%) than RM and RL (on average, less than 10%). The onset of laying (at least 10% laying rate) was shown, at different ages, according to the % ABW the breeds had reached: PP was the first, followed by ER, then RM, and RL was the last. At 42-53 weeks, the hen-day egg production ranged, on average, from 38 to 52%, according to the breeds; orthogonal contrasts on two-weekly data showed, at first age, increasing linear (ER) and quadratic (other groups) trends, and at second age, positive linear (ER, RM) and cubic (PP, RL) trends. Age (32 vs. 53 weeks) affected almost all the eggshell traits in PP and ER, whereas in RL, and especially RM, fewer traits changed. The age increased the yolk to albumen ratio (unchanged in PP). These results may be useful for the effective management of local purebred chickens, with the purpose to ensure the wellbeing of the hens and for supplying eggs of different quality throughout the year.

For laying hens, the immediate aftermath and healing period of a keel fracture (KF) is characterized by reduced ability to perform species-specific behavior, access resources, and pain. However, the longer-term impacts, once the fracture is completely healed, are less clear. As well as acute pain and behavioral changes, a negative experience can shape future responses to putatively threatening stimuli, raising future fear, and anxiety levels during husbandry-related events. We aimed to determine whether hens that had previously sustained keel bone fractures, but were now outside of the peak age range for new fractures, showed higher fear and anxiety levels compared to intact hens. We also determined if healed keel bone fractures were associated with reduced production, changes in behavior and resource use. One hundred and fifty hens with a palpation score of 1 ( "KF") and 150 hens with a palpation score of 0 (keel fracture free, "KFF") were selected from a commercial farm at 63 wk of age and housed in 6 groups (3 × KF and 3 × KFF). We compared production (hen weight and feed consumption, egg quantity, quality and weight, floor eggs, shell thickness, and weight) and home pen behavior (behaviors and transitional movements) in both groups. Finally, we measured the responses of KF (n = 75) and KFF (n = 75) during tonic immobility, novel arena, and novel object tests. KF and KFF hens did not differ in their responses to the tonic immobility, novel arena, and novel object tests, nor were there differences between the 2 groups in home pen behavior and transitional movements. KFF birds were lighter and laid eggs with less eggshell membrane compared KF birds, but no differences were found between KF and KFF in any other production measures. We found no evidence that healed KFs were associated with detrimental welfare effects in laying hens, but further work is required to determine the mechanisms and implications of the lower body weight and egg shell membrane., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Osteoporosis is a common degenerative metabolic bone disease in caged laying hens. Intensive egg production mobilizing large amounts of Ca from bone for eggshell formation, consequently leading to Ca deficiency, has been recognized as a critical factor causing osteoporosis in commercial laying hens. The aim of this study was to examine the effect of Ca deficiency on the function of the gut microbiota-bone axis and related egg production traits and bone health in laying hens. Twenty-four 48-week-old laying hens were fed a control diet (Control, 3.72%) or a low Ca diet (LC, 2.04%) for 60 d ( n  = 12). Compared to the Control hens, the LC hens had higher levels of alkaline phosphatase and tartrate resistant acid phosphatase ( P  < 0.05) with lower bone strength, eggshell thickness, and eggshell strength ( P  < 0.05). In addition, the LC hens had higher plasma estradiol concentrations, while having lower concentrations of interleukin-1 (IL-1) and IL-6. The LC hens also had a lower pH value in the ileum with an increased Ca retention. The principal co-ordinates analysis showed significantly separate cecal microbiota populations between the Control and LC hens. The Prevotellaceae&#95;UCG-001, Subdoligranulum, Peptococcus, and Eubacterium&#95;hallii&#95;group ( P  < 0.05) were higher, while the CHKC1001 and Sutterella ( P  < 0.05) were lower at the genus level in the LC hens. In addition, Prevotellaceae&#95;UCG-001, Subdoligranulum and Eubacterium&#95;hallii&#95;group had a negative correlation, while Sutterella was positively correlated with ileal pH values. The transcriptome analysis revealed that the low Ca diet caused 20 and 31 genes to be significantly up- and down-regulated, respectively. The gene expressions of cystic fibrosis transmembrane conductance regulator, solute carrier family 26 member 3 of the anion exchangers, and mitogen-activated protein kinase 12 of pro-inflammatory factors were lower in the LC birds, which was correlated with the lower ileal pH values. These results suggest that the hens with low Ca diet-induced osteoporosis have an increased intestinal Ca retention with a decreased ileal pH value, correlated with the changes in Prevotellaceae&#95;UCG-001, Subdoligranulum, and Eubacterium&#95;hallii&#95;group of beneficial genera. The results provide insights for further understanding and preventing osteoporosis in laying hens., Competing Interests: We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, and there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the content of this paper., (© 2023 The Authors. Publishing services by Elsevier B.V. on behalf of KeAi Communications Co. Ltd.)

A study was conducted to evaluate performance, eggshell quality, nutrient metabolizability, and incubation traits of laying breeder hens fed diets supplemented with an additive containing polyphenols and saponins of a proprietary blend from Quillaja saponaria and Yucca schidigera (QY) biomass. Hens were fed 4 feeds in 5 periods of 28 days each from 30 to 49 weeks of age. Experimental feeds were a Control diet; Control + virginiamycin (33 g/ton); Control + QY (250 g/ton) and Control + virginiamycin + QY. A total of 40 White Plymouth Rock and 44 Rhode Island Red breeder hens were allocated in individual cages using a completely randomized block design with 21 replicates. Performance parameters, evaluated per period, were egg production, egg weight, FCR, egg mass, and culled eggs. All eggs were collected in the last 4 days of each period to evaluate specific egg weight, percentage of albumen, yolk and shell, and Haugh unit as well as cuticle quality, shell strength, and shell thickness. At the end of the experiment, nutrient metabolizability assessment and four incubations were conducted. There were no interactions between diet and period in all evaluated responses ( P > 0.05). Experimental diets did not affect daily egg production, egg weight, and egg mass as well as Haugh unit, yolk and albumen percentage, and yolk color ( P > 0.05). However, hens fed Control + QY produced eggs with better shell strength, shell thickness, and cuticle quality than hens fed the Control ( P < 0.05). Hens fed Control + QY or Control + virginiamycin + QY had lower culled eggs, better FCR and higher egg specific weight, shell percentage, and yolk strength compared to breeder hens fed the Control ( P < 0.05). In general, hens fed QY achieved enhanced performance and egg quality compared to virginiamycin. In conclusion, laying breeder hens fed diets supplemented with Quillaja and Yucca additive, from 30 to 49 weeks of age, maintained their productive performance, had improved eggshell and cuticle quality and reduced culled, dirty and contaminated eggs., Competing Interests: Authors LG and SB were employed by Phibro Animal Health Corporation. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Souza, Adams, Agilar, Biselo, Mello, Gomez, Bonaspetti and Stefanello.)

Follicle selection and preovulatory hierarchy of hen ovaries were important stages of follicle development and crucially determining egg-laying performance. The selected follicles with a higher expression level of follicle-stimulating hormone receptor (FSHR) mRNA that facilitates response to FSH, and rapidly develops into preovulatory follicles with distinctive characteristics of granulosa cells (GCs) proliferation and differentiation. Identification of the key genes involved in these developmental events is helpful for elucidation of the molecular mechanism underlying egg-laying traits in chicken and other domestic fowl. Herein, the comparative transcriptomic analysis of ovarian prehierarchical follicles before selection (BSF), follicles at selection stage (ASF), and hierarchical follicles (HF) were implemented in the Jilin Black chicken (JB) and Lohmann Brown layer (LB) with the divergences in their egg-laying performance by RNA-sequencing. The results showed that nine deferentially expressed genes (DEGs), including STMN4, FABP3, ROBO2, RSPO4, and DMRT1 were revealed between follicles BSF and ASF; and seventeen DEGs, such as SLC6A15, SLITRK3, PRKG2 and TMC3 were mined between ASF and HF. These two group DEGs being co-expressed between BSF and ASF, and between ASF and HF were compared and substantiated in the JB and LB layers, respectively. Furthermore, 10 signaling pathways, such as cAMP signaling, PPAR signaling pathway, AMPK(Adenosine 5'-monophosphate (AMP)-activated protein kinase) pathway, and estrogen signaling pathway were also identified. Moreover, the roles of two representative candidates ROBO2 and PRKG2 genes presented as downregulated mRNA expression pattern in the transcriptomic profiles were further verified in vitro. The results demonstrated that downregulation of ROBO2 or PRKG2 significantly increased the expression levels of FSHR mRNA and protein with the boosted expression of CCND1, STAR, and BCL-2, whereas remarkably inhibited the expression of Caspase-3, consequently, brought about the decrease of GC apoptosis in the ovarian follicles, but increase of GC proliferation and differentiation serving as the hallmarks for follicle selection. It indicated that ROBO2 and PRKG2 may play indispensable roles in follicle selection and preovulatory hierarchy of hen ovaries separately. Our findings provided a comparative transcriptomic evidence for clarifying the molecular mechanism of the follicle development underlying egg-laying traits in chicken., (© The Author(s) 2023. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The aim of this study was to compare yield performance (from 39 to 50 weeks of age) and egg physical characteristics (at 50 weeks of age) of eight autochthonous chicken breeds of the Veneto region (Italy). Four white eggshell breeds, namely Padovana Camosciata (PA-C, chamois plumage), Padovana Dorata (PA-G, golden plumage), Polverara Bianca (PO-W, white plumage), and Polverara Nera (PO-B, black plumage), and four tinted eggshell breeds, namely Pepoi (PP), Ermellinata di Rovigo (ER), Robusta Maculata (RM), and Robusta Lionata (RL) from a conservation centre were considered in the trial. Significant differences (p < 0.05) among breeds were observed for yield performance and egg quality. From 39 to 50 weeks of age, the hen-day egg production was higher in PA-C and RM than in RL, and PO-W and ER were intermediate; PA-G, PO-B, and PP were the lowest. The hen-day egg production changed according to the age of the hens. From 39 to 42 weeks of age, ER showed the highest hen-day egg production and PA-G the lowest; from 47 to 50 weeks, PA-C, PO-W, and RM were the highest and PP the lowest. The tinted eggshell breeds, with the exception of PP, had higher egg weights than white eggshell breeds. PP egg weight was similar to PO-B. As regards the tinted eggshell breeds, RM eggs had the highest eggshell a* and b*, and PP the lowest. PA-C had the most spherical eggs, and PO-B and ER had the most ovoid eggs. PO-W and RM had the highest eggshell thickness and ER had the lowest. The highest eggshell ratio was observed for PO-W and PO-B, and the lowest for ER. The yolk-to-albumen ratio was higher in the white eggshell breeds than in PP, ER, and RL. ER had the highest Haugh units and PA-G the lowest. PO-W, PO-B, PA-C, PA-G, and ER had the lowest egg inclusions, and RL and RM the highest. Tinted eggshell eggs differed from white eggshell eggs by having higher meat spots. Results indicated that the eggs produced by the eight local chicken breeds differed according to the laying rate and a wide range of physical external and internal characteristics which allow the consumer to distinguish them for their genetic origin by the eggshell shape and colour, and to use them for different purposes to valorise poultry biodiversity.

The aims of the present study were to characterize egg composition and develop VIS-Near-infrared spectroscopy (VIS-NIR) models for its predictions in Italian local chicken breeds, namely Padovana Camosciata, Padovana Dorata, Polverara Bianca, Polverara Nera, Pepoi, Ermellinata di Rovigo, Robusta Maculata and Robusta Lionata. Hens were reared in a single conservation center under the same environmental and management conditions. A total of 200 samples (25 samples per breed, two eggs/sample) were analyzed for the composition of albumen and yolk. Prediction models for these traits were developed on both fresh and freeze-dried samples. Eggs of Polverara Nera and Polverara Bianca differed from eggs of the other breeds (p < 0.05) in terms of the greatest moisture content (90.06 ± 1.23% and 89.57 ± 1.31%, respectively) and the lowest protein content (8.34 ± 1.27% and 8.81 ± 1.27%) in the albumen on wet basis. As regards the yolk, Robusta Maculata and Robusta Lionata differed (p < 0.05) from the other breeds, having lower protein content (15.62 ± 1.13% and 15.21 ± 0.63%, respectively) and greater lipid content (34.11 ± 1.12% and 35.30 ± 0.98%) on wet basis. Eggs of Pepoi had greater cholesterol content (1406.39 ± 82.34 mg/100 g) on wet basis compared with Padovana Camosciata, Polverara Bianca and Robusta Maculata (p < 0.05). Spectral data were collected in reflectance mode in the VIS-NIR range (400 to 2500 nm) using DS2500 (Foss, Hillerød, Denmark) on fresh and freeze-dried samples. Models were developed through partial least-squares regression on untreated and pre-treated spectra independently for yolk and albumen, and using several combinations of scattering corrections and mathematical treatments. The predictive ability of the models developed for each compound was evaluated through the coefficient of determination (R2cv), standard error of prediction (SEcv) and the ratio of performance to deviation (RPDcv) in cross-validation. Prediction models performed better for freeze-dried than fresh albumen and yolk. In particular, for the albumen the performance of models using freeze-dried eggs was excellent (R2cv ≥ 0.91), and for yolk it was suitable for the prediction of protein content and dry matter. Good performances of prediction were observed in yolk for dry matter (R2cv = 0.85), lipids and cholesterol (R2cv = 0.74). Overall, the results support the potential of infrared technology to predict the composition of eggs from local hens. Prediction models for proteins, dry matter and lipids of freeze-dried yolk could be used for labelling purposes to promote local breeds through the valorization of nutritional aspects.

Hens are oviparous vertebrates and produce IgY antibodies, which is the main type of immunoglobulin in the egg yolk, and high concentrations can be obtained by using a simple method that does not require sophisticated equipment and reagents. The Polyethylene Glycol 6000 method allows the removal of lipids and the precipitation of IgY in two days with an approximated purity of around 80%, however during the original protocol other contaminant proteins can be precipitated. To overcome the issue of contamination with other proteins and extraction time, we optimized the previously method described by Pauly et al. (2011) by adding some changes that improved the aforementioned problems. • Our protocol is customized by the addition of one more filtration step or one more step with PEG 6000 at 3.5% to avoid the contamination with lipids. • Additionally, the changes in the type of agitation, centrifugation and the skip of dialysis make the method more accessible for all the laboratories. • In summary, these modifications serve to enhance the purity, reduce the time for IgY extraction from egg yolk and make it more accessible for every basic research laboratory., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Author(s).)

The importance of egg natural defences to prevent bacterial contamination and their relation with hen age in extended production cycles were evaluated. Egg-white from eggs of different hen age groups (up 100-weeks-old) and lines (Hy-Line white and brown) were inoculated with Gram-positive Staphylococcus aureus or Gram-negative Salmonella Typhimurium, ranging from 10 3 -10 6  CFU/mL. Our results show that concentrations of egg-white lysozyme and, particularly, ovotransferrin are important to modulate bacterial survival in a dose-dependent matter. Depending on protein concentration, their effect ranges from bactericidal to bacteriostatic, with a threshold for bacterial contamination that depends also on hen age and line. The concentrations of lysozyme and ovotransferrin increased with hen age (up to 2 and 22 w/w% of total protein, respectively), and eggs laid by older hens exhibited the greatest potential to prevent the growth of the highest Salmonella inoculum (10 6  CFU/mL). Salmonella-penetration experiments demonstrated that non-contaminated eggs display significantly higher concentrations of antimicrobial proteins. However, eggs from older hens needed a higher concentration of these proteins (>20% ovotransferrin) to prevent bacterial contamination, showing that antimicrobial protein concentrations in egg-whites was not the only factor influencing bacterial contamination. Finally, this study demonstrated that egg-white of eggs produced by old hens are less prone to contamination by Salmonella., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Embryo mortality rate, which can reach up to 40% in avian species, is a major issue for breeding. It is therefore important to identify new embryo development biomarkers for genetic selection to improve reproductive performances. We have recently shown that chemerin is expressed in the oviductal hen magnum, accumulates in egg white, is correlated with embryo survival and could thus be used as a molecular marker of embryo development. Eggs from seven hen breeds ( n = 70) were collected during five successive days at the end of the laying period. After weighing eggs, yolk and albumen, an egg white sample from each egg was collected and a blood sample was taken from each hen. Chemerin concentrations in albumen and blood samples were measured by a specific home made ELISA assay. Hen's plasma and egg's albumen chemerin levels were found to be correlated with reproductive parameters such as fecundity, fertility, embryo mortality, hatchability and laying rates. The inter-hen chemerin level variability in albumen was higher than intra-hen except for one breed (R+). We observed significantly different levels of chemerin in egg white between breeds. However, chemerin concentrations in egg white were not significantly associated to variations of hen plasma chemerin levels. Interestingly, we observed negative correlations between albumen chemerin concentrations and egg weight ( r = -0.43, p = 0.001), between albumen weight ( r = -0.40, p = 0.002), and between yolk weight ( r = -0.28, p = 0.03). We also showed negative correlations between egg white chemerin concentrations and fecundity ( r = -0.32, p = 0.011) and fertility ( r = -0.27, p = 0.04) whereas no significant correlation was observed with the laying rate. Taken together, these results suggest that egg white chemerin concentration might be a good biomarker for genetic selection for egg weight and fertility in hens, provided these data are confirmed on a larger scale., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Bernardi, Reverchon, Estienne, Baumard, Ramé, Brossaud, Combarnous, Froment and Dupont.)

In free-range and organic production systems, hens can make choices according to their needs and desires, which is in accordance with welfare definitions. Nonetheless, health and behavioral problems are also encountered in these systems. The aim of this article was to identify welfare challenges observed in these production systems in the EU and the most promising solutions to overcome these challenges. It is based on a review of published literature and research projects complemented by interviews with experts. We selected EU specific information for welfare problems, however, the selected literature regarding solutions is global. Free range use may increase the risk of infection by some bacteria, viruses and parasites. Preventive methods include avoiding contamination thanks to biosecurity measures and strengthening animals' natural defenses against these diseases which can be based on nutritional means with new diet components such as insect-derived products, probiotics and prebiotics. Phytotherapy and aromatherapy can be used as preventive and curative medicine and vaccines as alternatives to antibiotics and pesticides. Bone quality in pullets and hens prevents keel deviations and is favored by exercise in the outdoor range. Free range use also lead to higher exposure to variable weather conditions and predators, therefore shadow, fences and guard animals can be used to prevent heat stress and predation respectively. Granting a free range provides opportunities for the expression of many behaviors and yet many hens usually stay close to the house. Providing the birds with trees, shelters or attractive plants can increase range use. Small flock sizes, early experiences of enrichment and personality traits have also been found to enhance range use. Severe feather pecking can occur in free range production systems, although flocks using the outdoor area have better plumage than indoors. While many prevention strategies are facilitated in free range systems, the influence of genetics, prenatal and nutritional factors in free range hens still need to be investigated. This review provides information about practices that have been tested or still need to be explored and this information can be used by stakeholders and researchers to help them evaluate the applicability of these solutions for welfare improvement., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Bonnefous, Collin, Guilloteau, Guesdon, Filliat, Réhault-Godbert, Rodenburg, Tuyttens, Warin, Steenfeldt, Baldinger, Re, Ponzio, Zuliani, Venezia, Väre, Parrott, Walley, Niemi and Leterrier.)

Two hundred and twenty eggs from Transylvanian naked neck (TNN) chickens aged 30 wk were used to determine the effect of storage length on hatch window, chick quality, and organ development. Forty-four eggs (11 eggs in 4 replicates) were stored in cold room (16±1.5°C) in batches for 0, 3, 6, 9, and 12 d before setting in incubator. Data collected were subjected to one-way analysis of variance. Hatchability of set and fertile eggs declined (P < 0.001) progressively as duration of storage increased, however, storage for 6 and 9 d differed not from each other. For 50% hatch, there was a decrease of 30 min/d in hatching time by 3 d-stored eggs before fresh eggs, while eggs stored for 6, 9, and 12 d had delay of 70, 65, and 30 min/d, respectively. Hatching commenced earlier (P < 0.01) in eggs stored for 0 and 3 d than in prolonged storage of eggs for 6 to 12 d. Eggs stored for 12 d recorded shorter (P < 0.001) hatch window than those stored for 0 to 9d. Significantly (P < 0.01) more eggs hatched per hour during hatch window in fresh eggs (3.9%/h) than other storage durations (1.70-2.12 %/h). Fresh and 6 d eggs hatched into heavier (P < 0.05) chicks (37.2 and 37.5 g, respectively) than 12d (32.7g). Chicks from 9 and 12 d storage were shorter in length than those for 0 to 6 d. Toe (P < 0.01) and shank+toe (P < 0.001) were longer in 0 d chicks than others. Activity and appearance were poorer (P < 0.01) in 12 d chicks than other groups. The quality of eye was better (P < 0.05) in 0 to 6 d chicks than 12 d group, however, 9 d chicks were similar to 12 d. Larger membrane remnant were found in 6 to 12 d chicks (P < 0.001) than in 0 d chicks, though not different from 3 d group. Navel of chicks from eggs stored for 0, 3, and 6 d were more closed than in 9 d, though not different from 12 d. Yolk remnant was larger in chicks of 3, 9, and 12 d storage length than in fresh and 6 d storage length. Tona score was higher in 0 to 6 d chicks than in 9 and 12 d chicks. There was no (P > 0.05) effect of egg storage length on chick yield and relative weight of chick organs. It could be concluded that in order to ensure good hatchability and quality chicks, TNN eggs could be stored for 3 d, but not beyond 6 d without intervention such as prewarming of long-stored eggs., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

The effects of dietary carotenoids on egg yolk were investigated in this study. Forty Rhode Island Red (RR) and 40 Silky Fowl (SF) hens that were 60 weeks old were used. Hens of each breed were randomly divided into four dietary groups. One group was fed a basal diet (crude protein 17%, metabolizable energy 2800 kcal/kg) only, whereas the other groups received a specific additive, namely, paprika extract, marigold petal extract, or Paracoccus cell powder, in addition to the same basal diet. The color and carotenoid content of egg yolk and singlet oxygen quenching activity were measured after 4 weeks. The total carotenoid content, zeaxanthin content, and singlet oxygen quenching activity in the yolk differed significantly between breeds and between diets (two-way ANOVA). The lutein content in egg yolk was affected by breed and diet, as well as by the interaction between these two factors. Regarding the Roche Yolk Color Fan values, only the effect of diet was significant. In terms of objective egg yolk color, there was a significant difference in lightness and yellowness between breeds. The total carotenoid content was higher in SF than in RR in all the groups. Likewise, the levels of zeaxanthin and lutein in the yolk were higher in SF than in RR ( P <0.05). The results of the present study suggest that dietary carotenoids are effective feed additives for laying hens, especially SF, to improve the color and singlet oxygen quenching activity of egg yolk., (2022, Japan Poultry Science Association.)

Carotenoids, which are pigments known to have many health benefits, such as their antioxidant properties, are being researched for their potential as a feed additive for production animals. These pigments are found in varying quantities in different breeds of corn, and their impact on the chicken microbiome requires further investigation. This 35 d laying hen (Novagen White) feeding trial involved varying the levels and composition of carotenoids by changing the corn source: white (0.9 µg total carotinoids/g total diet), yellow (5.7 µg/g), and orange (24.9 µg/g). For each of the three corn diet treatments, 6 replicate cages were randomly assigned. The cecal microbial community composition of the hens was then studied by 16S rRNA gene amplicon sequencing. The composition of the cecal bacterial community, as determined by Bray-Curtis dissimilarity, was different (P < 0.05) in chickens fed the orange corn diet, compared to chickens on the white corn diet, but there was no statistical difference between animals fed yellow corn compared to the white or orange corn groups. There was no change in the alpha diversity between any of the groups. Within Lactobacillus, which is one of the most abundant genera, 2 amplicon sequence variants (ASVs) were decreased and one ASV was increased in the orange corn group compared to both the white and yellow corn groups. While previous studies showed that orange corn did not alter the community composition in broilers, it appears that orange corn based feed may alter the community composition of laying hens., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Responses to dietary energy and protein levels were compared between two egg-laying genotypes. Individually housed hens of a historic strain (HS) and a modern strain (MS) were compared. In Experiment 1 (Exp.1), four levels of true metabolizable energy, corrected for zero nitrogen retention (TME n ) and four levels of total lysine, were offered from 30 to 40 weeks of age. In Experiment 2 (Exp.2), three levels of apparent metabolizable energy, corrected for zero nitrogen retention (AME n ) and four levels of standardized ileal digestible lysine (SID Lys), were fed from 20 to 30 weeks of age. A randomized factorial block design (4 × 4 Exp.1 and 3 × 4 Exp.2) was applied. Energy utilization for egg output (EO) did not differ (NS), and both strains maintained a constant kJ intake (NS). The efficiency of SID Lys utilization for EO differed, with the MS being the more efficient ( p < 0.034). A single model could be used to predict feed intake, using BW, EO, AME n and SID Lys (r = 0.716). In conclusion, it is unlikely that the requirements of modern layer strains have increased. However, feeding programs should be adjusted for economic reasons and are dependent upon achievable feed intake under particular circumstances.

Eggshell translucency is a ubiquitous external eggshell quality problem caused by variations of eggshell ultrastructure or shell membrane. In previous studies, researchers have widely investigated this phenomenon with nutritional, environmental, and genetic perspectives in many breeds. However, most studies referring to phenotypic measurement of shell translucency have been performed using a relatively subjective two-, three-, or four-grading methods, which made it impossible to compare distribution of shell translucency among different breeds. In this study, we aimed to explore variations of translucent eggshell spots in different breeds and their distribution in blunt, middle, and sharp ends of eggshell using a relatively objective grayscale recognition method. We selected 45 eggs from each flock of pure lines, commercial strains, and Chinese local breeds (10 flocks, aged 60 to 70 wk), and stored them in a constant environment for 5 d. Then measured eggshell translucency using grayscale recognition method. Indicators of shell translucency included sum of spot areas on the whole eggshell (SUSA), sum area of the whole eggshell (SUSHA), RSS (ratio of SUSA to SUSHA), quantity of spots (QS), average spot area in eggshell (AAES), and diameter of spots in eggshell (DS). As results, in Hy-Line Brown, Brown-Egg Dwarf Layer, and Taihang (pink-shelled) breeds, phenotypic intensity of eggshell translucency was slight; in Rhode Island Red, Jingfen-1, and Dongxiang breeds, phenotypic intensity of eggshell translucency was relatively extensive; and in Jinghong-1, Hy-Line Sonia, White Leghorn, and Taihang (blue-shelled), phenotypic intensity of eggshell translucency was at an intermediate level. In general, the larger the RSS, the larger the QS, AAES, and DS. Of 3 ends for most breeds, eggshell translucency of blunt and sharp ends was usually greater than that of middle ends, and blunt ends seemed to have the most extensive eggshell translucency. Findings from this study could provide a reference for population selection to locate genes regulating shell translucency and to explore the physical structure mechanism for eggshell translucency formation., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Heat stress is one of the most detrimental environmental challenges affecting the biological process and the related production performance of farm animals, especially in poultry. Commercial laying hens have been bred (selected) for high egg production, resulting in increased sensitivity to heat stress due to breeding-linked metabolic heat production. In addition, laying hens are prone to heat stress due to their inadequate species-specific cooling mechanisms resulting in low heat tolerance. In addition, hens have no sweat glands and feathering covers almost their entire body to minimize body heat loss. The poultry industry and scientists are developing cooling methods to prevent or reduce heat stress-caused damage to chicken health, welfare, and economic losses. We have designed and tested a cooling system using perches, in which chilled water (10 °C) circulates through a conventional perch passing through the layer cages to offer the cooling potential to improve hen health, welfare, and performance during acute and chronic periods of heat stress (35 °C). This review summarizes the outcomes of a multi-year study using the designed cooled perch system. The results indicate that conducting heat from perching hens directly onto the cooled perch system efficiently reduces heat stress and related damage in laying hens. It provides a novel strategy: perches, one key furnishment in cage-free and enriched colony facilities, could be modified as cooling devices to improve thermal comfort for hens during hot seasons, especially in the tropical and subtropical regions.

Choline is an essential nutrient in laying hen diets and is needed for the formation of phosphatidylcholine (PC), that serves as a rich source of long chain (≥20 C) n-3 fatty acids (FA) in eggs. Methionine (Met) is the first limiting amino acid in layer hen diets and serves as a lipotropic agent with antioxidant properties. The objectives of the current study is based on the hypothesis that choline and Met supplementation will enhance egg PC and n-3 FA status, lipid stability, and production indices in layer hens fed flaxseed. Ninety-six, 40-wk-old laying hens (W-36 White Leghorns) were randomly allocated to 4 treatment groups, with 6 replicates containing four hens per cage. Hens were fed corn-soybean meal-based diet containing 0% flaxseed (Control), 15/100 g flaxseed (Flax), Flax+50% more methionine requirement for W-36 White Leghorns (Flax+Met), or Flax+0.15g/100g choline chloride (Cho) (Flax+Cho). All experimental diets were isocaloric and isonitrogenous and fed for a period of 120 d. Egg production and egg mass (g/hen/d) was higher for Flax+Met and Flax+Cho when compared to Flax and Control (P < 0.05). Egg weight was greater (P < 0.05) among hens fed the Control and Flax+Cho diets compared to Flax diet. Feeding flaxseed to hens led to over 6-fold increase in total n-3 FA. Choline supplementation increased egg α-tocopherol content (P < 0.05) while reducing lipid oxidation products measured as thiobarituric acid reactive substances in egg yolk (P < 0.05). Neither Met nor Cho had any impact on docosahexaenoic (22:6 n-3) acid concentration in eggs from hens fed flaxseed. However, addition of Met and Cho to layer diets increased docosapentaenoic acid (22:5 n-3) levels in eggs from hens fed flaxseed (P < 0.05). The PC content was lower in Control and Flax+Met (P < 0.05) when compared to Flax+Cho group. No difference was found in total lipid or phosphatidylethanolamine content of eggs (P > 0.05). The results from the current study suggest that n-3 FA content of egg yolk can be greatly increased by feeding flaxseed but reduced egg production. However, dietary Met and Cho can improve production performance in hens fed flaxseed-containing diets. Addition of Cho to flaxseed increased in egg weight, yolk α-tocopherol levels, PC content and oxidative stability of eggs when compared to hens fed flaxseed. Met and choline could be used in flaxseed (>15%) to increase egg production and egg mas., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

We have studied in detail the changes that occur in eggshell structure and composition during a production cycle in order to better understand the deterioration of eggshell quality with hen age (at 33, 45, and 67 wk). To study changes in eggshell ultrastructure and microstructure characteristics (mammillary density, palisade layer thickness, size, and orientation of calcite crystals) and the cuticle composition, we used complementary analytical techniques such optical and electron microscopy, X-ray diffraction, and infrared spectrometry. The marked decrease in eggshell breaking strength from 5.8 Kg at 33 wk to 4.4 Kg at 67 wk (25% reduction) could not be solely explained by the modest reduction in eggshell thickness (6-10% reduction) and seems to be associate to abrupt changes in eggshell ultra- and microstructure characteristics (i.e., decreased mammillary density; increased size of crystal units), occurring in older hens. Particularly, the decrease in mammillary density reduces the attachment points of the eggshell mineral to the membranes and therefore should negatively impact eggshell mechanical properties. Also, the observed increase in the calcite crystal size making the shell could also reduce the cohesion of crystals and eggshell resistance against impacts. Additionally, there was a decrease in the amount of cuticle and internal egg quality parameters (egg albumen height) with hen age that could have a negative impact in egg safety and quality., Competing Interests: DISCLOSURES All sources of funding are acknowledged in the manuscript, and authors have declared any direct financial benefits or conflict of interest that could result from publication. All appropriate ethics and other approvals were obtained for the research., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

The aim of the current study was to assess the impact of raspberry (RA), black currant (BC), and black chokeberry (CA) dried pomace, at 30 g per kg of feed, on performance, egg quality, white blood cells, heterophil:lymphocyte ratio, and lymphatic organs in Hy-Line laying hens. Hens fed a basal diet comprised the control group (CO). A total of 480, 42-week-old Hy-Line commercial hybrid laying hens were randomly divided into 4 equal groups differing in the diet: in control group (CO), the birds received a standard diet, in groups CH, BC and RA, the birds were fed with a standard diet enriched with 30 g/kg of dried fruit pomace: Black chokeberry, black currant, and raspberry, respectively. Differences in yolk color were observed between the CO and groups BC and RA, as well as between groups CA and BC (P < 0.001). The eggs of group RA were characterized by a lower weight (P = 0.001 and P = 0.002) and shell density (P = 0.025 and P = 0.002) in comparison to eggs from the control group CO. The lightest yolk color was observed in the eggs of group BC in comparison with the other groups (P = 0.006). The greatest yolk weight was observed for group CH, while the smallest was for group CO (P = 0.017). Laying hens in the groups BC and RA were characterized at wk 62 of age by a higher percentage of spleen in comparison with the control group CO (P = 0.018). In conclusion, the BC and RA diet resulted in a paler yolk color and was associated with slightly poorer shell quality parameters. On the other hand, the CH and BC diet had a positive impact on the immune system of the laying hens, as confirmed by the changes in the white blood cell smear and the higher spleen percentage. Therefore, the use of these by-products in the diets of poultry, i.e., dried pomace of black chokeberry and black currant may have a positive impact by improving the immunological status of laying hens., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Sulfamethoxazole-Trimethoprim residues in eggs can cause risks to human health. The most common cause of residues in eggs results from failure to meet an appropriate withdrawal interval. The aim of this study was to determine the quantity and duration of sulfamethoxazole-trimethoprim residues in eggs and evaluate the drug elimination parameters in egg components and whole egg to better estimate the withdrawal interval of sulfamethoxazole and trimethoprim following oral administration for 7 days at a purposed dosage regimen (time average 46 mg kg -1 day -1 for sulfamethoxazole, time average 25 mg kg -1 day -1 for trimethoprim). Residues of sulfamethoxazole and trimethoprim in albumen and yolk were analyzed by ultra-performance liquid chromatography mass spectrometry. A greater percentage of sulfamethoxazole was distributed into the albumen (91.53-96.74%) and a greater percentage of trimethoprim was distributed into yolk (63.92-77.36%) during treatment. The residues levels in whole egg declined below or reached the limit of quantification until 13 days for SMZ and TMP respectively. The withdrawal interval for SMZ and TMP were 43 days and 17 days respectively using the FDA tolerance method., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Qingyuan partridge chicken is an important indigenous chicken in China. In its breeding schemes, chickens are usually selected at the age of 105-day-old for five traits, including body weight (BW), shank length (SL), shank girth (SG), comb height (CH) and feather maturity (FM). At present, genetic parameters of the aforementioned traits have still not been studied in Qingyuan partridge chickens. The objectives of this study were the following: (1) to investigate whether the optimal statistical models of these traits need to consider maternal genetic and permanent environmental effects in late-feathering Qingyuan partridge hens, and (2) to estimate genetic parameters for these traits based on the optimal models. The numbers of records for BW, SL, SG, CH and FM were 13,721, 13,671, 13,670, 13,669 and 13,672, respectively. Variance components were estimated using average information-restricted maximum likelihood method, and the optimal model was determined based on Bayesian information criterion. More specifically, the optimal model for BW considered maternal genetic and permanent environmental effects in addition to direct additive genetic effect; SL, SG and FM considered direct and maternal genetic effects; and CH considered direct and maternal genetic effects, and the covariance between them. The direct heritabilities of these traits estimated using the optimal models were 0.21 ± 0.04, 0.30 ± 0.05, 0.40 ± 0.05, 0.59 ± 0.09 and 0.09 ± 0.04, respectively; the maternal heritabilities were 0.01 ± 0.04, 0.05 ± 0.05, 0.04 ± 0.05, 0.09 ± 0.09 and 0.03 ± 0.04, respectively. Maternal genetic effect evidently played an important part in FM and maternal heritability accounted for 30 per cent of total heritability. Furthermore, the direct and maternal genetic effects for CH were estimated to be negatively and moderately correlated (-0.51 ± 0.11). For all traits, neglecting existent maternal effects biased the estimation of direct heritability. Therefore, to implement optimum breeding strategies for improvement of these traits in Qingyuan partridge hens, maternal effects should be taken into consideration., (© 2020 Wiley-VCH GmbH.)

Little is known about early indicators of heat stress in bird erythrocytes. We investigated the effects of elevated temperatures on the morphology and cellular responses of avian erythrocytes. Hen red blood cells were subjected to 22-45 °C temperatures for 1 h and 4 h, then stained and examined by light microscopy to assess morphological alterations. Cell viability, cytotoxicity and caspases 3 and 7 activity also were investigated. We found that short-term exposure of hen blood to 43-45 °C caused morphological alterations and increased the activity of pro-apoptotic caspases 3 and 7; hemolytic cells also were found. Reduction of erythrocytes may be a consequence of direct disruption of the cell membrane, although apoptotic disintegration also may occur. Because changes in erythrocyte morphology were rapid, they may be useful indicators of thermal stress in birds.

Objective: Wheat bran (WB) was co-fermented with Aspergillus oryzae and phytase (Phy) to determine whether co-fermentation improve WB phosphorus and fiber utilization in Isa-brown layers., Methods: A total of 112 Isa brown layer were randomly divided into 7 treatments with 8 replicates per a treatment and 2 hens per a replicate. The treatments included basal diet (control), basal diet supplemented with 250 unit/kg Phy (control+Phy), diet with 10% WB (10% WB), diet with 5% WB and 250 unit/kg Phy (5% WB+Phy) diet with 10% WB and 250 unit/kg Phy (10% WB+Phy), diet with 5% fermented WB supplemented with molasses and phy (PCFWH) and 125 unit/kg Phy (5% PCFWH), and diet with 10% PCFWH (10% PCFWH). The intestinal microbial population, intestinal morphology, serum antioxidant enzyme activities, and excreta phosphorus content were assessed., Results: In PCFWH, spore counts, protease activity, xylanase activity, and ferulic acid were 8.50 log/g dry matter (DM), 190 unit/g DM, 120 unit/g DM, and 127 μg/g, respectively. Xylobiose and xylotriose were released in PCFWH, while they were not detectable in WB. Antioxidant capacity was also enhanced in PCFWH compared to WB. The 10% WB+Phy and 10% PCFWH groups produced higher egg mass, but hens fed 5% WB+Phy had the lowest amount of feed intake. Eggs from 10% PCFWH had better eggshell weight, eggshell strength, and eggshell thickness. Birds fed with 10% PCFWH also had higher serum superoxide dismutase and catalase activities. Compare to control, 10% PCFWH significantly reduced excreta phosphorus content., Conclusion: Diet inclusion of 10% PCFWH improved egg quality, antioxidant status, and excreta phosphorus content of laying hens.

Globally, the poultry industry is gaining significant importance among the agricultural and its allied sectors. However, heat stress was found to negatively affect the poultry production particularly in the tropical regions. This review is therefore an attempt to generate information pertaining to the impacts of heat stress on poultry production and its amelioration. Heat stress reduces the growth, reproductive performance, and egg production in poultry birds. The reduction in productive potential of poultry birds on exposure to heat stress may be attributed to the deviation of energy resources from production to adaptation pathway. There are different approaches pertaining to relieving the adverse impacts of heat stress on poultry production. These approaches can be broadly categorized under genetic, management, and nutritional strategies. These approaches may reduce the negative effects of heat stress and enhance the productive performance of poultry birds. The management strategies include appropriate shelter design, providing shade, using sprinklers, implementing cooling devices, and using fans and ventilation systems. The recommended floor space for mature birds weighing 1.7 kg is 0.06 m 2 /bird while it is 0.13 m 2 /bird for the birds weighing 3.5 kg with 27.8 kg/m 2 bird density in either case. The nutritional interventions comprise ration balancing and providing essential micronutrients to improve the productive and reproductive performance in poultry birds. Fat, antioxidants, yeast, and electrolyte supplementations are some of the most commonly used nutritional strategies to ensure optimum production in the poultry industry. Furthermore, providing adequate water supply and disease surveillance measures may help to ensure optimum meat and egg production in the birds. The advanced biotechnological tools may aid to identify suitable genetic markers in poultry birds which might help in developing new strains of higher thermo-tolerance by designing suitable breeding program involving marker-assisted selection. These strategies may help to optimize and sustain poultry production in the changing climate scenario.

Magnolol is a multifunctional plant polyphenol. To evaluate the effects of magnolol on laying hens in the late laying period, 360 (50-week-old) laying hens were randomly assigned to 4 dietary treatments: a non-supplemented control diet (C), and control diets supplemented with 100, 200, and 300 mg/kg of magnolol (M100, M200, and M300), respectively. Each treatment had 6 replicates with 15 hens per replicate. Results showed that dietary supplementation of 200 and 300 mg/kg of magnolol increased the laying rate and the M200 group had a lower feed conversion ratio (P < 0.05). Magnolol supplementation (200 and 300 mg/kg) could linearly increase albumen height and Haugh unit of fresh eggs in the late phase of the laying cycle (P < 0.01). And magnolol linearly alleviated the decline of the albumen height and Haugh unit of eggs stored for 14 d (P < 0.01). The total superoxide dismutase activity in the ovaries of M100 group was greater than that in the other treatments (P < 0.05). As dietary magnolol levels increased, villus height of jejunum and ileum linearly increased (P < 0.01). M200 and M300 groups had higher expression level of occludin in the ileum compared with group C (P < 0.01). The level of nitric oxide production and inducible nitric oxide synthase expression in the ileum of M200 group were lower than that in the C group (P < 0.05). In conclusion, dietary supplementation of 200 and 300 mg/kg magnolol can improve hen performance, albumen quality of fresh and storage eggs, and hepatic lipid metabolism in the late laying cycle. Also, magnolol has a good effect on increasing villi and improving the intestinal mucosal mechanical barrier function., (Copyright © 2020. Published by Elsevier Inc.)

Accumulating evidence has demonstrated the role of microRNAs (miRs) in the avian ovary. In this study, high-throughput transcriptome analyses were employed to study the differential miR expression profiles in the chicken ovary, aiming to reveal miR-targeting matrix metalloproteinase (MMP) expression during follicular growth, maturation, and atresia. Using tissues of chicken ovarian follicles at key steps of development (slow growing - white, the most recently recruited - small yellow, and preovulatory - F2) and regression (the third postovulatory), 14 small RNA (sRNA) libraries were constructed. The 25 most highly expressed known miRs were identified along with eight significantly differentially expressed (DE) miRs (gga-miR-let-7d, gga-miR-31-3p, gga-miR-138-1-3p, gga-miR-1552-5p, gga-miR-92-3p, gga-miR-31-5p, gga-miR-202-3p, and gga-miR-6648-3p) which were further examined by quantitative real time-PCR (qRT-PCR) in white, yellowish, small yellow, and atretic follicles as well as in the granulosa and theca layer of yellow preovulatory F3-F1 follicles (n = 6 hens). These miRs were mainly associated with four pathways: inhibition of MMPs, axonal guidance signaling, HIF1α signaling, and GP6 signaling. Four predicted target genes (i.e. MMP-16, ADAM10, COL4A2, and COL4A5) were examined by qRT-PCR and negatively correlated with DE miRs. The identified candidate miR:mRNA target pairs include gga-miR-31-5p or gga-miR-92-3p:MMP-16, gga-miR-31-5p or gga-miR-92-3p:ADAM10, let-7d:COL4A2, and gga-miR-138-1-3p:COL4A5 are potentially associated with MMP modulation in the hen ovary, mostly in the granulosa and theca cells of the largest preovulatory follicles. These results provide a novel insight to the role of miRs in follicle development by identifying a miR target network that is putatively engaged in remodeling of the extracellular matrix during ovarian follicle development in chickens., Competing Interests: Declaration of competing interest The authors declare no conflict of interest regarding the publication of this article., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The aim of this study was to investigate the effects of osteocalcin (OCN) on fatty liver hemorrhagic syndrome (FLHS) in aged laying hens. Thirty 68-week-old White Plymouth laying hens were randomly assigned into conventional single-bird cages, and the cages were randomly allocated into one of 3 treatments (n = 10): normal diet (ND + vehicle, ND + V), high-fat diet (HFD + vehicle, HFD + V), and HFD + OCN (3 μg/bird, 1 time/2 d, i.m.) for 40 d. At day 30, oral glucose tolerance tests (OGTT) and insulin tolerance tests (ITT) were performed. At the end of experiment, the hens were euthanized followed by blood collection. The plasma aspartate transaminase (AST), alkaline phosphatase (ALP), total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) were measured using an automatic biochemistry analyzer. Pathological changes in the liver were examined under both light and transmission electron microscopes. The plasma inflammatory factors including interleukin-1 (IL-1), IL-6, and tumor necrosis factor-alpha (TNF-α) were analyzed by ELISA, and the gene expressions of these inflammatory factors in the liver were analyzed by real-time PCR. The level of oxidative stress was evaluated using malondialdehyde (MDA) and glutathione peroxidase (GSH-Px) assay kits, respectively. The results showed that HFD + V hens had more severe liver hemorrhage and fibrosis than ND + V hens (P < 0.05). The ultramicrostructural examination showed that hepatocytes of HFD + V hens exhibited necrotic pyknosis showing great intracellular electron, mitochondrial swelling, shrunk nucleus, and absence of autolysosomes. Osteocalcin mitigated HFD + V-induced pathological changes in aged laying hens. High-fat diet + OCN hens had higher insulin sensitivity; lower liver concentrations of MDA (P = 0.12) but higher GSH-Px (P < 0.05); and lower blood TNF-α concentrations (P < 0.05) and mRNA expressions (P < 0.05) than HFD + V hens. These results suggest OCN functions in preventing the FLHS process in old laying hens through inhibiting excessive energy diet-induced metabolic disorder, oxidative stress, and related pathological damage., (Copyright © 2020. Published by Elsevier Inc.)

The effects of natto, a fermented soybean food, on transcript levels of hen peroxisome proliferator-activated receptor-γ (PPARG), PPARG coactivator-1α and -1β (PPARGC1A and PPARGC1B), and nuclear receptor corepressor 1 (NCOR1) were investigated using real-time polymerase chain reaction in white leghorn (Julia strain) hens. Twenty-one- and 34-week-old hens were fed a basic or 3% dried natto-supplemented diet for 8 weeks. In the 21- and 34-week-old hens fed the natto-supplemented diet, hepatic PPARGC1B and NCOR1 transcript levels and adipose and hepatic PPARG transcript levels were significantly lower, respectively, than those in the control group. Furthermore, 34- and 42-week-old hens were fed a basic diet supplemented with 3% of the protein/fiber-enriched fraction (PFB) or 0.6% of the fat-enriched fraction (FAT) of natto, respectively, for 8 weeks. Adipose PPARG transcript levels were higher in the FAT diet group and significantly lower in the PFB diet group than in the control group. However, both FAT and PFB diet groups showed significantly lower hepatic PPARG transcript levels than did the control group. These results suggest that dried natto influences the transcript levels of PPARG, PPARGC1B, and NCOR1, and the FAT and PFB of natto influence the adipose and hepatic PPARG transcript levels in hens., (© 2021 Japanese Society of Animal Science.)