Experience on postmortem minimally invasive tissue sampling to ascertain the cause of death determination in South African children: A case for implementing as standard of care.

Determining the death burden for prioritising public health interventions necessitates detailed data on the causal pathways to death. Postmortem minimally invasive tissue sampling (MITS), incorporating histology, molecular and microbial culture diagnostics, enhances cause-of-death attribution, particularly for infectious deaths. MITS proves a valid alternative to full diagnostic autopsies, especially in low- and middle-income countries. In Soweto, South Africa (SA), the Child Health and Mortality Prevention Surveillance (CHAMPS) programme has delineated over 1 000 child and stillbirth deaths since 2017. This SA CHAMPS site supports advocating for the use of postmortem MITS as routine practice, for more granular insights into under-5 mortality causes. This knowledge is crucial for SA's pursuit of Sustainable Development Goal 3.2, targeting reduced neonatal and under-5 mortality rates. This commentary explores the public health advantages and ethicolegal considerations surrounding implementing MITS as standard of care for stillbirths, neonatal and paediatric deaths in SA. Furthermore, based on the data from CHAMPS, we present three pragmatic algorithmic approaches to the wide array of testing options for cost-effectiveness and scalability of postmortem MITS in South African state facilities.

Examining the relationship between different naturally-occurring maxillary beak shapes and their ability to cause damage in commercial laying hens.

1. Using chicken models to avoid unnecessary harm, this study examined the relationship between naturally-occurring maxillary (top) beak shapes and their ability to cause pecking damage.2. A selection of 24 Lohmann Brown laying hens from a total population of 100 were sorted into two groups based on their maxillary beak shape, where 12 were classified as having sharp beaks (SB) and 12 as having blunt beaks (BB).3. All hens were recorded six times in a test pen which contained a chicken model (foam block covered with feathered chicken skin) and a video camera. During each test session, the number of feathers removed from the model, the change in skin and block weight (proxies for tissue damage) and the percentage of successful pecks (resulting in feather and/or tissue removal) were recorded.4. SB hens removed more feathers from the model and had a greater change in skin weight than BB hens. The mean number of pecks made at the model did not differ between the beak shape groups; however, SB hens had a greater percentage of successful pecks, resulting in feather and/or tissue removal, compared to BB hens.5. In conclusion, SB hens were more capable of removing feathers and causing damage. Birds performed more successful pecks resulting in feather and/or tissue removal as they gained experience pecking at the model.

An analysis of the maxillary beak shape variation between 2 pure layer lines and its relationship to the underlying premaxillary bone, feather cover, and mortality.

Beak shape varies considerably within and between intact-beak laying hens, and aspects of beak shape appear to be heritable. As an alternative to beak treatment (an effective method of reducing damage from severe feather pecking (SFP)), this variation could be used to genetically select hens whose beak shapes are less apt to cause damage. To be able to select certain phenotypes, the beak shape variation that exists within laying hen flocks must first be characterized. The objectives of this study were to 1) describe the maxillary beak shape variation in 2 pure White Leghorn layer lines with intact beaks using geometric morphometrics to analyze images, and 2) examine the beak shape's relationship to the premaxillary bone, feather cover, and mortality. A lateral head image was taken of each hen (n = 710), and 20 landmarks were placed along each image's dorsal and ventral margins of the maxillary beak. Landmark coordinates were standardized by Procrustes superimposition, and the covariation was analyzed by principal components analysis and multivariate regression. Feather cover was scored at 3 ages and mortality was monitored throughout the production cycle. Three principal components (PCs) explained 83% of the maxillary beak shape variation and the first PC partially separated the 2 lines. Maxillary beak shapes ranged from long and narrow with pointed tips to short and wide with more curved tips. Moderate correlations were found between the maxillary beak and premaxillary bone shape (rs = 0.44) and size (rs = 0.52). Line A hens had better feather cover than Line B at all ages. Line A hens also had less total and cannibalism-related mortality than Line B (10.7 and 0.4% vs. 16.7 and 2.4%, respectively). Beak shape may be one factor contributing to the observed differences in feather cover and mortality. The results suggest that distinct maxillary beak phenotypes within each line could be selected to help reduce SFP damage and improve bird welfare.

Development of an in vivo radiographic method with potential for use in improving bone quality and the welfare of laying hens through genetic selection.

1. Genetic selection for bone quality can improve this, as it is heritable. A method was established using digital X-ray which took around 40 s in total and gave an image that allowed quantification of bone density from many appendicular bones.2. The tibiotarsus measurement of bone density on the live hen across the different experiments had correlations with post-mortem whole bone radiographic density from 0.62 to 0.7, similar to that between density and material properties for example. Differences between groups of hens, where calcium and phosphorus in the diet were manipulated, were detected within 3 weeks of treatment using live hen measurement (P < 0.001, n = 24).3. In a gage analysis, 'hen' explained more than 86% of the variance, demonstrating the ability to observe clear differences between hens. The effect of different operators' analysis on the contribution to variance was very low as was the repeated measurement of the same hen.4. The measurement of bone density on the live hen described in this paper represented major progress to a usable method for genetic selection to improve bone strength in laying hens. The method has the potential to reduce the number of animals needed to test nutritional and management interventions to improve bone health.

Determining the variation in premaxillary and dentary bone morphology that may underlie beak shape between two pure layer lines.

Beak treatment is an effective method of reducing the damage inflicted by severe feather pecking (SFP) but there is significant pressure to eliminate these treatments and rely solely on alternative strategies. Substantial variation in beak shape exists within non-beak treated layer flocks and beak shape appears to be heritable. There is the potential to use this pre-existing variation and genetically select for hens whose beak shapes are less apt to cause damage during SFP. To do this, we must first understand the range of phenotypes that exist for both the external beak shape and the bones that provide its structure. The objective of this study was to determine the variation in premaxillary (within the top beak) and dentary (within the bottom beak) bone morphology that exists in 2 non-beak treated pure White Leghorn layer lines using geometric morphometrics to analyze radiographs. Lateral head radiographs were taken of 825 hens and the premaxillary and dentary bones were landmarked. Landmark coordinates were standardized by Procrustes superimposition and the covariation was analyzed by principal components analysis and multivariate regression using Geomorph (an R package). Three principal components (PCs) explained 85% of total premaxillary bone shape variation and showed that the shape ranged from long and narrow with pointed bone tips to short and wide with more curved tips. Two PCs explained 81% of total dentary bone shape variation. PC1 described the dentary bone length and width and PC2 explained the angle between the bone tip and its articular process. For both bones, shape was significantly associated with bone size and differed significantly between the two lines. Bone size accounted for 42% of the total shape variation for both bones. Together, the results showed a range of phenotypic variation in premaxillary and dentary bone shape, which in turn may influence beak shape. These bone phenotypes will guide further quantitative genetic and behavioral analyses that will help identify which beaks shapes cause the least damage when birds engage in SFP.

Review: What are the challenges facing the table egg industry in the next decades and what can be done to address them?

There has been a strong consumer demand to take welfare into account in animal production, including table eggs. This is particularly true in Europe and North America but increasingly around the world. We review the main demands that are facing the egg industry driven by economic, societal and sustainability goals. We describe solutions already delivered by research and those that will be needed for the future. Already table egg consumption patterns have seen a major shift from cage to non-cage production systems because of societal pressures. These often feature free-range and organic production. These changes likely signal the future direction for the layer sector with the acceleration of the conversion of cage to barn and aviary systems with outdoor access. This can come with unintended consequences from bone fracture to increased disease exposure, all requiring solutions. In the near future, the laying period of hens will be routinely extended to improve the economics and environmental footprint of production. Many flocks already produce close to 500 eggs per hens in a lifetime, reducing the number of replacement layers and improving the economics and sustainability. It will be a challenge for scientists to optimize the genetics and the production systems to maintain the health of these hens. A major ethical issue for the egg industry is the culling of male day-old chicks of layer breeds as the meat of the males cannot be easily marketed. Much research has and will be devoted to alternatives. Another solution is elimination of male embryos prior to hatching by in ovo sexing approaches. The race to find a sustainable solution to early stage sex determination is on. Methods based on sex chromosomes, sexually dimorphic compounds and spectral properties of eggs containing male or female embryos, are being researched and are reviewed in this article. Other proposed solutions include the use of dual-purpose strains, where the males are bred to produce meat and the females to produce eggs. The dual-purpose strains are less efficient and do not compete economically in the meat or egg market; however, as consumer awareness increases viable markets are emerging. These priorities are the response to economic, environmental, ethical and consumer pressures that are already having a strong impact on the egg industry. They will continue to evolve in the next decade and if supported by a strong research and development effort, a more efficient and ethical egg-laying industry should emerge.

Comparative analysis of the morphology, chemistry and structure of the tibiotarsus, humerus and keel bones in laying hens.

1. Bone properties are adapted to their specific functions in the animal, so various types of bones develop different characteristics depending on their location in the skeleton.2. The aim of this research was to compare the chemical composition, mineral characteristics and structural organisation in tibiotarsus, humerus and keel bones as representatives of hen skeletal mineralisation. Complementary analytical techniques, such as X-ray radiography, optical and electron microscopy, thermogravimetry and 2D X-ray diffraction, were used for characterisation.3. The humerus had a thinner cortex and cortical bone mineral had higher crystallinity and a greater degree of crystal orientation than the tibiotarsus. The humerus generally lacks medullary bone although, when present, it has a higher mineral content than seen in the tibiotarsus. These differences were attributed to the different forces that stimulate bone formation and remodelling.4. The keel cortical bone had a lower degree of mineralisation than the tibiotarsus or humerus. Its degree of mineralisation decreased from the cranial to the distal end of the bone. This gradient may affect keel mechanical properties, making it more prone to deformation and fractures.5. Data from studying different bones in laying hens can help to understand mineralisation as well as finding solutions to prevent osteoporosis-related fractures.

Dietary betaine reduces plasma homocysteine concentrations and improves bone strength in laying hens.

1. This study tested the hypothesis that the methyl-donor properties of betaine could reduce homocysteine concentrations, which has been recognised in a previous genetics study to be linked to bone quality. This was combined with phytase treatment, as phosphorus is critical for bone mineralisation.2. Using a 2 × 2 factorial arrangement, a total of 1920 Lohmann LSL-lite chickens housed as 24 replicates of 20 chickens were fed one of four diets containing dietary betaine (0 or 1000 mg/kg) and phytase (300 or 1000 FTU/kg) from one day old until end-of-lay. Blood and bone samples were collected at 45 and 70 weeks of age.3. Hens fed betaine had lower plasma homocysteine level (P < 0.05), higher tibia breaking strength (P < 0.05) and higher tibia bone density (P < 0.05).4. Egg production and quality was excellent throughout the study and were not affected by the dietary treatments.5. The addition of dietary betaine was successful at reducing plasma homocysteine concentrations and improving bone strength in laying hens, which could be used as an intervention to alleviate welfare concerns.

Sex differences in basal hypothalamic anorectic and orexigenic gene expression and the effect of quantitative and qualitative food restriction.

BACKGROUND: Research into energy balance and growth has infrequently considered genetic sex, yet there is sexual dimorphism for growth across the animal kingdom. We test the hypothesis that in the chicken, there is a sex difference in arcuate nucleus neuropeptide gene expression, since previous research indicates hypothalamic AGRP expression is correlated with growth potential and that males grow faster than females. Because growth has been heavily selected in some chicken lines, food restriction is necessary to improve reproductive performance and welfare, but this increases hunger. Dietary dilution has been proposed to ameliorate this undesirable effect. We aimed to distinguish the effects of gut fullness from nutritional feedback on hypothalamic gene expression and its interaction with sex. METHODS: Twelve-week-old male and female fast-growing chickens were either released from restriction and fed ad libitum or a restricted diet plus 15% w/w ispaghula husk, a non-nutritive bulking agent, for 2 days. A control group remained on quantitative restriction. Hypothalamic arcuate nucleus neuropeptides were measured using real-time PCR. To confirm observed sex differences, the experiment was repeated using only ad libitum and restricted fed fast-growing chickens and in a genetically distinct breed of ad libitum fed male and female chickens. Linear mixed models (Genstat 18) were used for statistical analysis with transformation where appropriate. RESULTS: There were pronounced sex differences: expression of the orexigenic genes AGRP (P < 0.001) and NPY (P < 0.002) was higher in males of the fast-growing strain. In genetically distinct chickens, males had higher AGRP mRNA (P = 0.002) expression than females, suggesting sex difference was not restricted to a fast-growing strain. AGRP (P < 0.001) expression was significantly decreased in ad libitum fed birds but was high and indistinguishable between birds on a quantitative versus qualitative restricted diet. Inversely, gene expression of the anorectic genes POMC and CART was significantly higher in ad libitum fed birds but no consistent sex differences were observed. CONCLUSION: Expression of orexigenic peptides in the avian hypothalamus are significantly different between sexes. This could be useful starting point of investigating further if AGRP is an indicator of growth potential. Results also demonstrate that gut fill alone does not reduce orexigenic gene expression.

Validated quantitative trait loci for eggshell quality in experimental and commercial laying hens.

Compromised eggshell quality causes considerable economic losses for the egg industry. Breeding for improved eggshell quality has been very challenging. Eggshell quality is a trait that would greatly benefit from marker-assisted selection, which would allow the selection of sires for their direct contribution to the trait and would also allow implementation of measurements integrating a number of shell parameters that are difficult to measure. In this study, we selected the most promising autosomal quantitative trait loci (QTL) affecting eggshell quality on chromosomes 2, 3, 6 and 14 from earlier experiments and we extended the F2 population to include 1599 F2 females. The study was repeated on two commercial populations: Lohmann Tierzucht Rhode Island Red line (n = 692 females) and a Hy-Line White Plymouth Rock line (n = 290 progeny tested males). We analyzed the selected autosomal QTL regions on the three populations with SNP markers at 4-13 SNPs/Mb density. QTL for eggshell quality were replicated on all studied regions in the F2 population. New QTL were detected for eggshell color on chromosomes 3 and 6. Marker associations with eggshell quality traits were validated in the tested commercial lines on chromosomes 2, 3 and 6, thus paving the way for marker-assisted selection for improved eggshell quality.

Development of Azeliragon, an Oral Small Molecule Antagonist of the Receptor for Advanced Glycation Endproducts, for the Potential Slowing of Loss of Cognition in Mild Alzheimer's Disease.

Increasing evidence supports the role of the Receptor for Advanced Glycation Endproducts (RAGE) in the pathology of Alzheimer's disease. Azeliragon (TTP488) is an orally bioavailable small molecule inhibitor of RAGE in Phase 3 development as a potential treatment to slow disease progression in patients mild AD. Preclinical studies in animal models of AD (tgAPPSwedish/London) have shown azeliragon to decrease Aß plaque deposition; reduce total Aß brain concentration while increasing plasma Aß levels; decreases sAPPß while increasing sAPPα; reduce levels of inflammatory cytokines; and slow cognitive decline and improve cerebral blood flow. In the Phase 2b study, 18-months treatment in patients with mild-to-moderate AD indicated a baseline to endpoint change in ADAS-cog of 3.1 points in favor of drug. A greater magnitude of effect was evident in the sub-group of patients with mild AD (MMSE 21-26) with a baseline to endpoint change of 4 points on the ADAS-cog in favor of azeliragon and a 1 point change in CDR-sb in favor of drug. Azeliragon 5 mg/day delayed time to cognitive deterioration (7-point change in ADAS-cog from baseline, logrank p=0.0149). Based on promising results from the Phase 2b study, a Phase 3 registration program (STEADFAST) is being conducted under a Special Protocol Assessment from FDA. The ongoing Phase 3 program, if successful may demonstrate azeliragon can slow cognitive decline in mild AD patients.

Influence of physical activity on tibial bone material properties in laying hens.

Laying hens develop a type of osteoporosis that arises from a loss of structural bone, resulting in high incidence of fractures. In this study, a comparison of bone material properties was made for lines of hens created by divergent selection to have high and low bone strength and housed in either individual cages, with restricted mobility, or in an aviary system, with opportunity for increased mobility. Improvement of bone biomechanics in the high line hens and in aviary housing was mainly due to increased bone mass, thicker cortical bone and more medullary bone. However, bone material properties such as cortical and medullary bone mineral composition and crystallinity as well as collagen maturity did not differ between lines. However, bone material properties of birds from the different type of housing were markedly different. The cortical bone in aviary birds had a lower degree of mineralization and bone mineral was less mature and less organized than in caged birds. These differences can be explained by increased bone turnover rates due to the higher physical activity of aviary birds that stimulates bone formation and bone remodeling. Multivariate statistical analyses shows that both cortical and medullary bone contribute to breaking strengthThe cortical thickness was the single most important contributor while its degree of mineralization and porosity had a smaller contribution. Bone properties had poorer correlations with mechanical properties in cage birds than in aviary birds presumably due to the greater number of structural defects of cortical bone in cage birds.

Diurnal and photoperiodic changes in thyrotrophin-stimulating hormone ß expression and associated regulation of deiodinase enzymes (DIO2, DIO3) in the female juvenile chicken hypothalamus.

Increased thyrotrophin-stimulating hormone ß (TSHß) expression in the pars tuberalis is assumed to be an early step in the neuroendocrine mechanism transducing photoperiodic information. The present study aimed to determine the relationship between long-photoperiod (LP) and diurnal TSHß gene expression in the juvenile chicken by comparing LP-photostimulated birds with groups kept on a short photoperiod (SP) for 1 or 12 days. TSHß expression increased by 3- and 23-fold after 1 and 12 days of LP-photostimulation both during the day and at night. Under both SP and LP conditions, TSHß expression was between 3- and 14-fold higher at night than in the day, suggesting that TSHß expression cycles in a diurnal pattern irrespective of photoperiod. The ratio of DIO2/3 was decreased on LPs, consequent to changes in DIO3 expression, although there was no evidence of any diurnal effect on DIO2 or DIO3 expression. Plasma prolactin concentrations revealed both an effect of LPs and time-of-day. Thus, TSHß expression changes in a dynamic fashion both diurnally and in response to photoperiod.

Rapid Mass Spectrometry Imaging to Assess the Biochemical Profile of Pituitary Tissue for Potential Intraoperative Usage.

Pituitary adenomas are relatively common intracranial neoplasms that are frequently treated with surgical resection. Rapid visualization of pituitary tissue remains a challenge as current techniques either produce little to no information on hormone-secreting function or are too slow to practically aid in intraoperative or even perioperative decision-making. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) represents a powerful method by which molecular maps of tissue samples can be created, yielding a two-dimensional representation of the expression patterns of small molecules and proteins from biologic samples. In this chapter, we review the use of MALDI MSI, its application to the characterization of the pituitary gland, and its potential applications for guiding the management of pituitary adenomas.

Increasing persistency in lay and stabilising egg quality in longer laying cycles. What are the challenges?

In the past 50 years, selection starting initially at the breed level and then using quantitative genetics coupled with a sophisticated breeding pyramid, has resulted in a very productive hybrid for a variety of traits associated with egg production. One major trait currently being developed further is persistency of lay and the concept of the "long life" layer. Persistency in lay however cannot be achieved without due consideration of how to sustain egg quality and the health and welfare of the birds in longer laying cycles. These multiple goals require knowledge and consideration of the bird's physiology, nutritional requirements, which vary depending on age and management system, reproductive status and choice of the selection criteria applied. The recent advent of molecular genetics offers considerable hope that these multiple elements can be balanced for the good of all in the industry including the hens. The "long life" layer, which will be capable of producing 500 eggs in a laying cycle of 100 weeks, is therefore on the horizon, bringing with it the benefits of a more efficient utilisation of diminishing resources, including land, water, raw materials for feed as well as a reduction in waste, and an overall reduced carbon footprint.

Identification and characterisation of alternative transcriptional variants of PDGFRL in two lines of commercial poultry.

The platelet-derived growth factor (PDGF) family of genes and their receptors are involved in angiogenesis and steroid hormone production. A putative member of the family, platelet-derived growth factor receptor-like (PDGFRL), has been implicated in steroid-based feedback mechanisms within the chicken reproductive system. Three potential variants of PDGFRL were identified in the chicken, supported by in silico prediction and EST sequencing. The three potential transcripts have been further verified and the 5' terminal regions sequenced in this research. The sum of expression of all three transcripts in broiler breeders (the parents of broiler chickens) has been shown to be consistent with total expression of the gene. However, cumulative expression of the three transcripts in a range of tissues in egg layers was significantly short of total expression, indicating the existence of potential additional variants. Two additional variants were subsequently identified in egg layer cerebellum tissue and the 5' terminal regions sequenced. Sequence analysis of the three initial variants suggests that only one variant, which was the most abundant in broiler breeders and the majority of egg layer tissues, had a functional signal peptide. Although 5' RACE identified two additional transcripts in egg layers, the most likely protein translations indicated that these variants possessed no functional signal peptide, suggesting that, if they have a function, it is not a traditional one.

Hypothalamic Agouti-Related Peptide mRNA is Elevated During Natural and Stress-Induced Anorexia.

As part of their natural lives, animals can undergo periods of voluntarily reduced food intake and body weight (i.e. animal anorexias) that are beneficial for survival or breeding, such as during territorial behaviour, hibernation, migration and incubation of eggs. For incubation, a change in the defended level of body weight or 'sliding set point' appears to be involved, although the neural mechanisms reponsible for this are unknown. We investigated how neuropeptide gene expression in the arcuate nucleus of the domestic chicken responded to a 60-70% voluntary reduction in food intake measured both after incubation and after an environmental stressor involving transfer to unfamiliar housing. We hypothesised that gene expression would not change in these circumstances because the reduced food intake and body weight represented a defended level in birds with free access to food. Unexpectedly, we observed increased gene expression of the orexigenic peptide agouti-related peptide (AgRP) in both incubating and transferred animals compared to controls. Also pro-opiomelanocortin (POMC) mRNA was higher in incubating hens and significantly increased 6 days after exposure to the stressor. Conversely expression of neuropeptide Y and cocaine- and amphetamine-regulated transcript gene was unchanged in both experimental situations. We conclude that AgRP expression remains sensitive to the level of energy stores during natural anorexias, which is of adaptive advantage, although its normal orexigenic effects are over-ridden by inhibitory signals. In the case of stress-induced anorexia, increased POMC may contribute to this inhibitory role, whereas, for incubation, reduced feeding may also be associated with increased expression in the hypothalamus of the anorexigenic peptide vasoactive intestinal peptide.

Genome-wide association study for egg production and quality in layer chickens.

Discovery of genes with large effects on economically important traits has for many years been of interest to breeders. The development of SNP panels which cover the whole genome with high density and, more importantly, that can be genotyped on large numbers of individuals at relatively low cost, has opened new opportunities for genome-wide association studies (GWAS). The objective of this study was to find genomic regions associated with egg production and quality traits in layers using analysis methods developed for the purpose of whole genome prediction. Genotypes on over 4500 birds and phenotypes on over 13,000 hens from eight generations of a brown egg layer line were used. Birds were genotyped with a custom 42K Illumina SNP chip. Recorded traits included two egg production and 11 egg quality traits (puncture score, albumen height, yolk weight and shell colour) at early and late stages of production, as well as body weight and age at first egg. Egg weight was previously analysed by Wolc et al. (2012). The Bayesian whole genome prediction model--BayesB (Meuwissen et al. 2001) was used to locate 1 Mb regions that were most strongly associated with each trait. The posterior probability of a 1 Mb window contributing to genetic variation was used as the criterion for suggesting the presence of a quantitative trait locus (QTL) in that window. Depending upon the trait, from 1 to 7 significant (posterior probability >0.9) 1 Mb regions were found. The largest QTL, a region explaining 32% of genetic variance, was found on chr4 at 78 Mb for body weight but had pleiotropic effects on other traits. For the other traits, the largest effects were much smaller, explaining <7% of genetic variance, with regions on chromosomes 2, 12 and 17 explaining above 5% of genetic variance for albumen height, shell colour and egg production, respectively. In total, 45 of 1043 1 Mb windows were estimated to have a non-zero effect with posterior probability > 0.9 for one or more traits.

Hypothalamic agouti-related protein expression is affected by both acute and chronic experience of food restriction and re-feeding in chickens.

The central melanocortin system is conserved across vertebrates. However, in birds, little is known about how energy balance influences orexigenic agouti-related protein (AGRP) and anorexigenic pro-opiomelanocortin (POMC) expression, despite the fact that commercial food restriction is critical to the efficient production of poultry meat. To enable contrasts to be made, in broiler-breeder chickens, between levels of food restriction, between birds with the same body weight but different feeding experience, and between birds moved from restricted feeding to ad lib. feeding for different periods, five groups of hens were established between 6 and 12 weeks of age with different combinations of food restriction and release from restriction. AGRP and neuropeptide Y expression in the basal hypothalamus was significantly increased by chronic restriction but only AGRP mRNA levels reflected recent feeding experience: hens at the same body weight that had recently been on ad lib. feeding showed lower expression than restricted birds. AGRP expression also distinguished between hens released from restriction to ad lib. feeding for different periods. By contrast, POMC and cocaine- and amphetamine-regulated transcript mRNA levels were not different. These results showed that AGRP mRNA not only reflected differences between a bird's weight and its potential weight or set point, but also discriminated between differing feeding histories of birds at the same body weight. Therefore, AGRP expression potentially provides an integrated measure of food intake experience and an objective tool to assess a bird's perception of satiety in feeding regimes for improved poultry welfare.