Bone quality and composition are influenced by egg production, layer line, and oestradiol-17ß in laying hens.

Keel bone fractures are a serious animal welfare problem in laying hens. The aim of the current study was to assess the influence of egg production, oestradiol-17ß, and selection for high laying performance on bone quality. Hens of two layer lines differing in laying performance (WLA: 320 eggs per year, G11: 200 eggs per year) were allocated to four treatment groups. Group S received a deslorelin acetate implant that suppressed egg production. Group E received an implant with the sexual steroid oestradiol-17ß. Group SE received both implants and group C did not receive any implant. In the 63rd week of age, composition and characteristics of the tibiotarsi were assessed using histological analysis, three-point bending test, thermogravimetric analysis, infrared spectroscopy, and two-dimensional X-ray diffraction, respectively. Non-egg-laying hens showed a higher total bone area and a higher relative amount of cortical bone compared to egg-laying hens. Hens of layer line G11 showed a higher relative amount of medullary bone and a higher degree of mineralization of the cortical bone compared to hens of layer line WLA. These differences in bone composition may explain different susceptibility to keel bone fractures in non-egg-laying compared to egg-laying hens as well as in hens of layer lines differing in laying performance. The effect of exogenous oestradiol-17ß on bone parameters varied between the layer lines indicating a genetic influence on bone physiology and the way it can be modulated by hormone substitution.

Comparison of cortisol concentrations in plasma and saliva in dairy cattle following ACTH stimulation.

The experiments described in this research communication compared cortisol concentrations in plasma and saliva samples collected from dairy cattle before and after an adrenocorticotropic hormone (ACTH) application. For that purpose, blood and saliva samples were collected from five dairy cows at determined time intervals before (490 min and down to 0 min) and after (10 min and up to 500 min) an ACTH application. Mean baseline cortisol concentrations were greater in plasma compared to saliva. The relative increases and decreases in plasma and salivary cortisol concentrations following ACTH were similar. After ACTH, we observed an increase in cortisol concentrations in plasma after 10 min and in saliva after 20 min. The time of peak concentrations after ACTH were reached at 70 and 80 min for plasma and saliva, respectively. After peak concentrations, values steadily declined and returned to baseline values at 169 ± 15 min in plasma and 170 ± 14 min in saliva. Ratios between salivary and plasma cortisol concentrations were on average 0.09 and did not change substantially during the ACTH challenge. There was a strong positive relationship between salivary and plasma cortisol concentrations. These results indicate that salivary cortisol concentrations can be a good indicator of ACTH-induced plasma cortisol concentrations in dairy cattle.

Japanese quails (Coturnix Japonica) show keel bone damage during the laying period-a radiography study.

Keel bone damage is an important welfare issue in laying hens and can occur with a high prevalence of up to 100% of hens within one flock. Affected hens suffer from pain. Although multiple factors contribute to the prevalence and severity of keel bone damage, selection for high laying performance appears to play a key role. With up to 300 eggs/year, Japanese quails show a high laying performance, too, and, thus, may also show keel bone damage. However, to our knowledge, there are no scientific results on keel bone damage in Japanese quails to date. Therefore, the aim of this study was to assess whether keel bone fractures and deviations occur in Japanese quails and to obtain more detailed information about the development of their keel bone during the production cycle. A group of 51 female quails were radiographed at 8, 10, 15, 19, and 23 weeks of age. The X-rays were used to detect fractures and deviations and to measure the lateral surface area, length, and radiographic density of the keel bone. In addition, the length of the caudal cartilaginous part of the keel bone was measured to learn more about the progress of ossification. At 23 weeks of age, quails were euthanized and their macerated keel bones assessed for fractures and deviations. Both keel bone deviations and keel bone fractures were detected in the Japanese quails. In the 23rd week of age, 82% of the quails had a deviated keel bone as assessed after maceration. Furthermore, there was a decrease in radiographic density, lateral surface area, and length of the keel bone between weeks of age 8 and 19. This could indicate a general loss of bone substance and/or demineralization of the keel bone. Our study shows that keel bone damage is not only a problem in laying hens but also affects female Japanese quails.

A tagged visual analog scale is a reliable method to assess keel bone deviations in laying hens from radiographs.

Laying hens often suffer from keel bone damage (KBD) that includes pathologies with different etiologies, like diverse forms of fractures and deviations. Since KBD is a problem in all countries and housing systems, methods for the assessment of deviations are urgently needed. Comparisons between genetic lines and between studies are important to detect underlying mechanisms. Field researchers often use palpation as a low-cost and feasible technique for the assessment of KBD. In contrast to palpation, radiography is effective and highly precise at least in detecting keel bone fractures. The aim of this study was to: i) develop a scoring system to assess keel bone deviations from radiographs, ii) to assess inter- and intra-observer reliability of this scoring system, and iii) to investigate whether fractures and deviations of the keel are correlated. In total, 192 hens were used for the investigation. Digital radiographs were taken and evaluated for all hens after slaughter. We developed a tagged visual analog scale with two extreme images as anchors and four intermediate tags, resulting in six images representing the range from "no deviation" to "highly deviated" on a 10 cm line. Eleven participants scored 50 radiographs of keels with varying degree of severity, whereas five images were scored twice to assess intra-observer reliability. Intraclass correlation coefficient for inter-observer reliability was 0.979 with a confidence interval of 0.968 < ICC < 0.987 (F49,268 = 54.2, p < 0.0001). Intraclass correlation coefficient for intra-observer reliability was 0.831 with a confidence interval of 0.727 < ICC < 0.898 (F54,55 = 10.8, p < 0.0001). Individual intra-observer reliability ranged from 0.6 to 0.949. The Spearman correlation showed a strong positive correlation of fractures and deviations (s roh= 0.803, p < 0.001). The tagged visual analog scale could be a reliable instrument for the scoring of keel bone deviations. Our results support the assumption that the majority of highly deviated keels suffer from fractures as well. Further research is needed to investigate the correlation of palpation scores with the evaluation on radiographs.

Estradiol-17ß Is Influenced by Age, Housing System, and Laying Performance in Genetically Divergent Laying Hens (Gallus gallus f.d.).

The estrogen estradiol-17ß is known as one of the major gonadal steroid hormones with different functions in reproduction. In this study we analyzed estradiol-17ß concentration in laying hens of four pure bred chicken laying lines at four different time intervals of the laying period (17th-19th week of age, 33rd-35th week of age, 49th-51st week of age, and 72nd week of age). The high performing white egg (WLA) and brown egg (BLA) layer lines as well as the low performing white (R11) and brown (L68) layer lines were kept in both single cages and a floor housing system. We investigated whether there were differences in estradiol -17ß concentrations between lines at different ages that could be related to selection for high egg production or phylogenetic origin of the animals, and whether there was an influence of housing conditions on estradiol-17ß. Estradiol-17ß concentrations differed between high and low performing layer lines at all time intervals studied. High performing hens showed higher estradiol-17ß concentrations compared to low performing hens. In all lines, highest estradiol-17ß concentration was measured at their 49th to their 51st week of age, whereas the peak of laying intensity was observed at their 33rd to their 35th week of age. Additionally, hens with fewer opportunities for activity housed in cages showed higher estradiol-17ß concentrations than hens kept in a floor housing system with more movement possibilities. We could show that laying performance is strongly linked with estradiol -17ß concentration. This concentration changes during laying period and is also influenced by the housing system.

Productive performance, perching behavior, keel bone and other health aspects in dual-purpose compared to conventional laying hens.

Several alternatives to avoid killing male day-old chicks are available. One of these alternatives is to keep dual-purpose chicken strains. The aim of this study was to compare dual-purpose hens (Lohmann Dual, LD) with conventional laying hens (Lohmann Tradition, LT) in terms of performance, animal welfare parameters such as keel bone state and foot pad dermatitis, and perching behavior. We expected a generally equal or even better performance of the dual-purpose hens except for laying performance. Four hundred female day-old chicks were housed in 6 pens (3 pens per strain) and reared until 54 wk of age. Each pen offered a littered area, elevated slatted manure pit, elevated wooden frame with perches or grids and nest boxes on the manure pit. The wooden frame was alternately equipped with perches or grids. The elevated manure pit as well as the elevated structure was accessible via ramp. Productive performance parameters like mortality, total number of eggs and body weight were assessed periodically. In week 49, 132 hens (66 hens per strain) were randomly selected for radiography of the keel bone and assessment of plumage and foot pad state. Perching behavior was analyzed via scan sampling during rearing and laying period, respectively. Statistical analyzes were done with Linear Mixed Effect Model and General Linear Mixed Model. LD had a higher radiographic density than LT hens (P = 0.0016), other keel bone parameters (fracture score, P = 0.36; deformation, P = 0.83) showed no differences. The vast majority of fractures occurring in both strains were located in the caudal part of the keel bone. During the laying period, usage of elevated structures was higher with grids compared to perches (P < 0.001) and in LD compared to LT (P = 0.01). Some animal welfare problems were less frequent in LD compared to LT hens while other problems did not differ between the 2 strains or were even more frequent in LD hens. Grids may be more suitable as resting area than perches and may possibly help to decrease the prevalence of keel bone damage.

Video Endoscopy-Guided Intrabronchial Spray Inoculation of Mycobacterium bovis in Goats and Comparative Assessment of Lung Lesions With Various Imaging Methods.

Bovine tuberculosis (bTB) not only poses a zoonotic threat to humans but also has a significant economic impact on livestock production in many areas of the world. Effective vaccines for humans, livestock, and wildlife are highly desirable to control tuberculosis. Suitable large animal models are indispensable for meaningful assessment of vaccine candidates. Here, we describe the refinement of an animal model for bTB in goats. Intrabronchial inoculation procedure via video-guided endoscopy in anesthetized animals, collection of lungs after intratracheal fixation in situ, and imaging of lungs by computed tomography (CT) were established in three goats using barium sulfate as surrogate inoculum. For subsequent infection experiments, four goats were infected with 4.7 × 102 colony-forming units of M. bovis by intrabronchial inoculation using video-guided endoscopy with spray catheters. Defined amounts of inoculum were deposited at five sites per lung. Four age-matched goats were mock-inoculated. None of the goats developed clinical signs until they were euthanized 5 months post infection, but simultaneous skin testing confirmed bTB infection in all goats inoculated with M. bovis. In tissues collected at necropsy, M. bovis was consistently re-isolated from granulomas in lymph nodes, draining the lungs of all the goats infected with M. bovis. Further dissemination was observed in one goat only. Pulmonary lesions were quantified by CT and digital 2D radiography (DR). CT revealed mineralized lesions in all the infected goats ranging from <5 mm to >10 mm in diameter. Small lesions <5 mm predominated. The DR failed to detect small lesions and to determine the exact location of lesions because of overlapping of pulmonary lobes. Relative volume of pulmonary lesions was low in three but high in one goat that also had extensive cavitation. CT lesions could be correlated to gross pathologic findings and histologic granuloma types in representative pulmonary lobes. In conclusion, video-guided intrabronchial inoculation with spray catheters, mimicking the natural way of infection, resulted in pulmonary infection of goats with M. bovis. CT, but not DR, presented as a highly sensitive method to quantify the extent of pulmonary lesions. This goat model of TB may serve as a model for testing TB vaccine efficacy.

Influence of Age and Phylogenetic Background on Blood Parameters Associated With Bone Metabolism in Laying Hens.

The high laying performance of today's laying hens places enormous demands on their mineral metabolism. While up-to-date data are rare, the present study aimed to describe blood parameters associated with egg laying and bone metabolism during the pre-laying period, in the course of the laying period and the daily egg laying cycle. Ten to 15 laying hens of two high-performing, phylogenetically divergent lines (BLA: brown-egg layer; WLA: white-egg layer), kept in single cages were blood sampled at 17, 25, 29, 49, and 69 weeks of age. Sampling was made at 6 a.m., 10 a.m., 2 p.m. and, with the exception of week 17, 6 p.m. Blood samples were analyzed for concentrations of total and ionized calcium, inorganic phosphate (PO4), markers of bone formation (osteocalcin) and resorption [carboxyterminal crosslinked telopeptide of type I collagen (CTX-I)], 25-hydroxycholecalciferol (25(OH)D3) and estradiol-17ß. In the pre-laying period (17 week), the estradiol-17ß level calculated for WLA was more than twice as high as the level calculated for BLA, while no significant difference could be observed in the laying period (25 to 69 weeks). BLA hens had significantly higher total calcium concentrations at 49 weeks of age as well as up to twice as high levels of osteocalcin and 25(OH)D3 than WLA at any time of the day from 25 to 69 weeks of age. While osteocalcin, CTX-I and 25(OH)D3 concentrations were significantly higher before the onset of lay, total calcium and estradiol-17ß levels significantly increased from 17 to 69 weeks of age. In contrast, PO4 values varied only slightly during the experimental period and ionized calcium was highest at 17 and 49 weeks of age and lowest around peak production (29 week). In the course of the daily egg laying cycle blood concentrations clearly reflected the stage of egg formation. Our results provide up-to-date data of bone- and egg laying-associated blood parameters of two contemporary purebred layer lines over the course of the pre- and egg-laying period and the daily egg laying cycle. Differences between brown- and white-egg layers raise questions, whether phylogenetic background determines their efficiency to cope with high calcium demands relating to egg production.

Investigation of the Morphology of Adrenal Glands in Hens Kept in Two Different Housing Systems-A Pilot Study.

It is difficult to objectively assess the chronic effects of housing systems on livestock and particularly on laying hens. However, this seems to be important in the context of animal welfare. Therefore, we conducted the present study in order to compare the effect of two different housing conditions, single cage (SC) and floor pen (FP), on the morphology of the adrenal gland. A higher amount of interrenal cells, which secrete stress hormones, can lead to a difference in the relation of adrenal and interrenal cells, which could be interpreted as an indication of chronic stress. For this purpose, adrenal glands were extracted, prepared, stained and examined by microscopy, and total area of the cut, total area of interrenal cells and total area of adrenal cells were measured. As a result, all laying hens had a higher percentage of interrenal cells than adrenal cells (FP: interrenal cells/adrenal cells = 78.37%/21.63%; SC: 80.00%/20.00%). The median of adrenal-interrenal ratio did not differ significantly (FP = 0.2503, SC = 0.2499), while the variation of the ratio between laying hens in FP and SC showed a slight tendency of a higher ratio in adrenal glands of FP (p < 0.0870). Body weight and adrenal-interrenal ratio were significantly negatively correlated in laying hens in FP (rS = -0.943, p < 0.0048) but not in SC (rS = -0.162, p = 0.7283). There was no significant correlation between body weight and total cell area for interrenal cells or adrenal cells. Body weight was significantly lower for laying hens kept in SC than for laying hens kept in FP (p < 0.0001). Due to the present results, it can be concluded that keeping laying hens in single cages can have a negative effect on body weight.

Smaller brains in laying hens: New insights into the influence of pure breeding and housing conditions on brain size and brain composition.

During domestication, many different chicken breeds have been developed that show many alterations compared with their wild ancestors and large variability in parameters such as body size, coloring, behavior, and even brain morphology. Among the breeds, one can differentiate between commercial and noncommercial strains, and commercial strains do not usually show variability as high as noncommercial breeds but exhibit a high production rate of eggs (or meat). The breeding of high-performing laying hens, including the housing conditions of hens, is often a focus of concern for animal welfare, and to date, little is known about the correlation between housing conditions and artificial selection on brain structure. Based on an allometric approach, we compared the relative brain sizes of 2 inbred strains of laying hens (WLA and R11) with those of 7 other noncommercial chicken breeds. In addition, we examined the brain composition of laying hens and analyzed the relative sizes of the telencephalon, hippocampus, tectum opticum, and cerebellum. Half of WLA and R11 lines were kept in floor-housing systems, and the other half were kept in a single cage-housing system. Both strains of laying hens showed significantly smaller brains than the other chicken breeds. In addition, there was a significant difference between WLA and R11 hens, with R11 hens having larger brains. There was no difference in the relative brain sizes of floor-housed and cage-housed hens. WLA and R11 hens did not differ in their brain composition, but floor-housed hens showed a significantly larger cerebellum than cage-housed hens. Apparently, pure breeding over a long time and strong artificial selection for a high production of eggs is accompanied by (unintentional) selection for smaller brains. Further studies may also reveal differences in brain composition and the influence of housing conditions on brain composition.

The Role of Egg Production in the Etiology of Keel Bone Damage in Laying Hens.

Keel bone fractures and deviations belong to the most severe animal welfare problems in laying hens and are influenced by several factors such as husbandry system and genetic background. It is likely that egg production also influences keel bone health due to the high demand of calcium for the eggshell, which is, in part, taken from the skeleton. The high estrogen plasma concentration, which is linked to the high laying performance, may also affect the keel bone as sexual steroids have been shown to influence bone health. The aim of this study was to investigate the relationship between egg production, genetically determined high laying performance, estradiol-17ß concentration, and keel bone characteristics. Two hundred hens of two layer lines differing in laying performance (WLA: high performing; G11: low performing) were divided into four treatment groups: Group S received an implant containing a GnRH agonist that suppressed egg production, group E received an implant containing the sexual steroid estradiol-17ß, group SE received both implants, and group C were kept as control hens. Between the 12th and the 62nd weeks of age, the keel bone of all hens was radiographed and estradiol-17ß plasma concentration was assessed at regular intervals. Non-egg laying hens showed a lower risk of keel bone fracture and a higher radiographic density compared to egg laying hens. Exogenous estradiol-17ß was associated with a moderately higher risk of fracture within egg laying but with a lower risk of fracture and a higher radiographic density within non-egg laying hens. The high performing layer line WLA showed a significantly higher fracture risk but also a higher radiographic density compared to the low performing layer line G11. In contrast, neither the risk nor the severity of deviations were unambiguously influenced by egg production or layer line. We assume that within a layer line, there is a strong association between egg production and keel bone fractures, and, possibly, bone mineral density, but not between egg production and deviations. Moreover, our results confirm that genetic background influences fracture prevalence and indicate that the selection for high laying performance may negatively influence keel bone health.

Explanations for keel bone fractures in laying hens: are there explanations in addition to elevated egg production?

The current article served to provide the most up-to-date information regarding the causes of keel bone fracture. Although elevated and sustained egg production is likely a major contributing factor toward fractures, new information resulting from the development of novel methodologies suggests complementary causes that should be investigated. We identified 4 broad areas that could explain variation and increased fractures independent of or complementing elevated and sustained egg production: the age at first egg, late ossification of the keel, predisposing bone diseases, and inactivity leading to poor bone health. We also specified several topics that future research should target, which include continued efforts to link egg production and bone health, examination of noncommercial aves and traditional breeds, manipulating of age at first egg, a detailed histological and structural analysis of the keel, assessment of prefracture bone condition, and the relationship between individual activity patterns and bone health.

Bone Health or Performance? Adaptation Response of Genetically Divergent Chicken Layer Lines to a Nutritive Calcium Depletion.

In modern laying hybrids, calcium (Ca) homeostasis is immensely challenged by daily eggshell calcification. However, excessive mobilization of Ca from bones may lead to osteoporosis, which then manifests in a high incidence of poor bone quality. The aim of this study was to characterize the hens' adaptation response to an alternating dietary Ca restriction. The animal model consisted of four purebred layer lines, differing in laying performance (high vs. moderately performing lines) and phylogenetic origin (white- vs. brown-egg lines). According to the resource allocation theory, hens selected for high egg production were assumed to show a different response pattern to cope with this nutritive challenge compared to moderately performing lines. Data collected included egg number, egg quality traits, body weight and bone characteristics. The Ca depletion led to a temporary drop in egg production and shell quality and a loss of bone stability due to Ca mobilization. The white-egg lines response was more pronounced, whereas the brown-egg lines were less sensitive towards reduced Ca supply. Our study shows that the hens' responsiveness to coping with a nutritive Ca depletion is not ultimately linked to genetic selection for increased egg production but rather to phylogenetic origin.

Effects of intra-hippocampal D-AP5 injections on one trial passive avoidance learning in adult laying hens (Gallus gallus domesticus).

Domestic chickens are an established model organism for studies on learning and memory. Commonly, the chicks are used as subjects in several different learning tests, including one trial learning tests. However, for adult laying hens no such one trial learning tests have been established. In particular, there is no test established which focuses on the role of the hippocampus, a brain region, which is often critically involved in learning and memory consolidation. In this study we investigated the inhibitory effects of intra-hippocampal D-AP5 injections on a specific one trial passive avoidance learning test in adult laying hens (Gallus gallus domesticus). We used a step down avoidance learning paradigm (SDA) which is frequently used in rodents. Intra-hippocampal injections of D-AP5 impaired the learning abilities of adult laying hens compared to sham-injected control subjects. Thus, the experiments revealed that the hippocampus is critically involved in learning the inhibitory SDA task. Our results further indicate that the step down avoidance paradigm is suitable to examine learning and memory processes in adult laying hens.

Influence of a Sustained Release Deslorelin Acetate Implant on Reproductive Physiology and Associated Traits in Laying Hens.

The aim of this study was to develop an animal model with non-laying hens which would allow for investigation of the relationship between egg production and common diseases in hens. A total of 40 Lohmann Selected Leghorn hens were kept for 20 weeks in a floor housing system in two groups: group "Adult" (21 weeks of age) and group "Juvenile" (14 weeks of age). In each group, 10 hens were administered a 4.7 mg sustained release deslorelin acetate implant subcutaneously; in group "Adult" after, in group "Juvenile" before the onset of lay. In both groups, the remaining hens served as control hens. An examination of each hen was performed weekly, including ultrasonography to check for ovarian follicles, analysis of estradiol-17ß plasma concentration, and assessment of comb size. Digital radiographs of the keel bone were taken in experimental weeks 7 and 15. No follicles were detected on the ovary of treated hens for a certain time period which varied between individuals (between 8 weeks and until the end of the experiment). Estradiol-17ß concentrations were significantly higher in control hens. The comb was significantly smaller in treated hens. A lower prevalence of keel bone damage (group "Adult") and foot pad dermatitis (FPD) (both groups) was found in treated compared to control hens. These results show that a model with laying and non-laying hens can be achieved by administering a deslorelin acetate implant. Furthermore, they indicate a relationship between egg production and keel bone damage as well as FPD.

Radiographic examination of keel bone damage in living laying hens of different strains kept in two housing systems.

A high prevalence of deviations and fractures of the keel bone is a widespread welfare problem in laying hens. The aim of this study was to experimentally investigate this multifactorial problem throughout the laying period and to compare the prevalence and severity in different layer lines and different housing systems. High performing white (WLA) and brown (BLA) pure bred layer lines and low performing white (R11, G11) and brown layer lines (L68) were kept in both single cages and a floor housing system. A total of 97 hens (19 or 20 from each line, respectively) were repeatedly radiographed in the 35th, 51st and 72nd week of age. Fracture prevalence increased with age (p<0.001). The proportion of deviated keel bone area increased only for caged BLA, WLA and R11 hens (p<0.05) and was significantly higher for caged WLA and R11 hens compared to floor-housed WLA and R11 hens in the 72nd week of age (p<0.05). In the 72nd week of age hens in the floor housing system showed significantly more fractures than hens kept in cages (p<0.05). Prevalence of keel bone deviations was significantly higher in the white layer line R11 but significantly lower in the white layer line G11 compared to both brown layer lines and WLA (p<0.05). Brown layers showed significantly more fractures than white layers (p<0.05) in the 51st and 72nd week of age. Within the brown layers there was a significantly lower prevalence of deviations (p<0.05) and fractures (p<0.05) in the low performing (L68) compared to the high performing line (BLA). Our results show a different development of keel bone damage in caged compared to floor-housed hens under experimental conditions. Additionally, they indicate genetic effects on keel bone damage.