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

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

Part of this work was supported by the Spanish Ministerio de Ciencia e Innovacion [grant number CGL2015-64683-P].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.Instituto de Salud Carlos III Spanish Government CGL2015-64683-

Der Einfluss von Legetätigkeit, Genetik, Alter und Haltungssystem auf die Entstehung von Brustbeinschäden bei Legehennen

The keel bone is the prominent ventral part of the sternum in birds where the flight muscles attach. It is fractured or deviated, i.e., deformed, in up to 97 % or 83 % of laying hens within one flock, respectively. Both symptoms are often summarized to the term “keel bone damage” (KBD). Keel bone fractures and possibly also deviations are likely to cause pain and impair the mobility of affected hens. For these reasons, KBD is considered to be one of the most severe animal welfare problems in the egg production industry. The etiology of KBD is not yet fully understood but it is widely defined as a multifactorial disorder. There are external factors such as housing system and nutrition as well as internal factors such as genetic background and age influencing the prevalence of KBD. However, there is no agreement about the direction of these effects and knowledge about the extent to which each of these factors contributes to the etiology of fractures and deviations is lacking. Another internal factor that may influence the keel bone is egg production. There is a high demand of calcium for the eggshell. To meet this demand, female birds possess a special kind of woven bone which is located in the medullary cavity of some bones. It is suggested that once the hen comes into lay, osteoblasts change their function from forming structural bone to forming medullary bone which leads to a decrease in the amount of structural bone and, thus, in bone strength. It is further suggested that these mechanisms are mediated by estrogens. However, the role of egg production and estrogens in KBD has never been investigated into detail. The aim of the present work was to get a better insight into the etiology of KBD. A special focus was put on comparing the external factor housing system with the internal factors genetic background, age, egg production, and estradiol-17β. To that aim, three studies were carried out. In the first study, a method to assess keel bone fractures and deviations in living hens was established using radiography. Furthermore, hens of five layer strains differing in phylogenetic background (brown versus white layer lines) as well as laying performance (high versus low performing) were kept in two different housing systems (single cages versus floor housing) and repeatedly radiographed. Brown layer lines showed more keel bone fractures while the severity of keel bone deviations tended to be higher in white layer lines. Within the brown layers, the high performing layer line showed more keel bone fractures and deviations compared to the low performing layer line. More fractures were found in the floor housing system whereas keel bone deviations were more severe in cages within some of the layer lines. Fracture prevalence increased with age. The presented radiographic examination of the keel bone allowed to clearly differentiate between fractures and deviations and to assess the severity of the latter. It was further shown to be a suitable and quick method for longitudinal studies on KBD. The aim of the second study was to establish an animal model with non-egg laying hens which could further be used to assess the influence of egg production on different traits in laying hens. 40 hens were kept in a floor housing system. Ten hens received a sustained release implant containing the gonadotropin releasing hormone (GnRH) agonist deslorelin acetate before and ten hens after the onset of lay. The remaining 20 hens were kept as control hens. The implant inhibited egg laying activity in all hens that were treated after the onset of lay and protracted the onset of lay in all hens that were treated before. However, duration of effectiveness was relatively short and showed that a new implant should be administered after approximately twelve weeks in order to constantly inhibit egg laying activity. Furthermore, estradiol-17β plasma concentration was decreased in treated hens. All hens of this study were also radiographed twice. Egg laying control hens showed significantly more keel bone fractures and more severe keel bone deviations compared to non-egg laying hens within the group that was treated after the onset of lay. Furthermore, severity of keel bone deviations increased with age in this group. The results of the first two studies were used for the third study whose aim was to assess the influence of egg production and estradiol-17β on KBD. A total of 200 laying hens of two strains differing in laying performance were kept in a floor housing system. Half of each layer line was administered a deslorelin acetate implant every 90 days and, thus, did not lay eggs. Part of these hens as well as of the egg laying hens was further given an implant with estradiol-17β. All hens were repeatedly radiographed and fracture prevalence as well as prevalence and severity of deviations were compared between the four treatment groups and both layer lines. Furthermore, radiographic density of the keel bone was assessed. The risk of keel bone fracture was much lower in non-egg laying compared to egg laying hens while no effect of egg laying activity on keel bone deviations was found. Radiographic density of the keel bone was higher in aged non-egg laying hens compared to aged egg laying hens. Treatment with exogenous estradiol only showed a relatively small effect on keel bone fracture risk within egg laying hens and no effect on deviations or radiographic density. The high performing layer line showed a higher risk of keel bone fracture than the low performing layer line but layer lines did not differ in terms of keel bone deviations. Taken together, a method to assess keel bone fractures, deviations and radiographic density in a longitudinal study as well as a model with non-egg laying hens have been established and can be used in further studies. Different risk factors have been found for keel bone fractures and deviations indicating that these are two different and independent phenomena and that it is very important to clearly differentiate between them. Both external and internal factors have been found to contribute to the etiology of keel bone fractures and deviations. Part of the keel bone fractures seem to be caused by collisions with housing equipment. However, the very large difference in risk of keel bone fracture between egg laying and non-egg laying hens clearly indicates that there is a fundamental weakness of the keel bone in laying hens caused by egg production which makes it very susceptible to fractures. Findings about the higher prevalence of keel bone fractures in high compared to low performing layer lines support this assumption. It is, thus, necessary to figure out which are the differences in bone structure and composition between egg laying and non-egg laying hens in order to find solutions against this huge animal welfare problem.Der Begriff „Brustbeinschäden“ umfasst Frakturen und Deformationen der Carina sterni, die bei flugfähigen Vögeln sehr ausgeprägt ist und als Ansatzfläche für die Flugmuskulatur dient. Brustbeinschäden kommen bei Legehennen sehr häufig vor: Bis zu 97 % der Hennen einer Herde können von Frakturen und bis zu 83 % der Hennen von Deformationen betroffen sein. Da Brustbeinfrakturen und möglicherweise auch -deformationen mit hoher Wahrscheinlichkeit schmerzhaft sind und die Bewegungsfähigkeit der betroffenen Tiere beeinträchtigen, werden Brustbeinschäden als eines der größten Tierschutzprobleme in der Legehennenhaltung betrachtet. Die Ursachen von Brustbeinfrakturen und -deformationen sind noch nicht ausreichend geklärt. Es wird davon ausgegangen, dass es sich um ein multifaktorielles Krankheitsbild handelt, auf dessen Entstehung sowohl exogene Faktoren wie das Haltungssystem und die Fütterung als auch endogene Faktoren wie die Genetik und das Alter der Hennen Einfluss haben. Jedoch widersprechen sich die vorhandenen Studien teilweise bezüglich der Richtung der Effekte und auch das Ausmaß, in welchem die einzelnen Faktoren jeweils Einfluss auf die Entstehung von Brustbeinschäden nehmen, ist unbekannt. Ein weiterer endogener Faktor, der eine Rolle in der Entwicklung von Brustbeinschäden spielen könnte, ist die Legetätigkeit. Legehennen haben durch die Eischalenbildung einen sehr hohen Calciumbedarf. Dieser Bedarf wird teilweise durch Bereitstellung von Calcium aus dem Skelett, v.a. dem medullären Knochen, gedeckt. Dies ist ein spezielles geflechtartiges Knochengewebe, das bei weiblichen Vögeln in der Markhöhle einiger Knochen zu finden ist. Es besteht die Theorie, dass die Osteoblasten mit Legebeginn der Henne nur noch medullären und keinen kortikalen sowie trabekulären Knochen mehr bilden, was zu einer Abnahme der Knochenstabilität führt. Es wird weiterhin vermutet, dass diese Mechanismen durch Östrogene gesteuert werden. Jedoch gibt es bis heute keine Studien, die den Einfluss der Legetätigkeit und von Östrogenen auf die Brustbeingesundheit untersucht haben. Das Ziel der vorliegenden Arbeit war es, einen tieferen Einblick in die Ätiologie von Brustbeinschäden zu gewinnen. Hierbei wurden insbesondere der exogene Faktor Haltungssystem und die endogenen Faktoren Genetik, Alter, Legetätigkeit und 17β-Östradiol untersucht. Zu diesem Zweck wurden drei verschiedene Studien durchgeführt. Das Ziel der ersten Studie war es, eine Röntgenmethode zu entwickeln, die eine Verlaufsuntersuchung von Brustbeinfrakturen und -deformationen sowie eine zuverlässige Unterscheidung dieser beiden Symptome und eine Vermessung von Deformationen erlaubt. Des Weiteren wurden fünf Legelinien und zwei Haltungssysteme miteinander verglichen. Die Legelinien unterschieden sich in ihrer phylogenetischen Herkunft (Braun- und Weißleger) sowie in ihrer Legeleistung (Hoch- und Minderleistung). Von jeder Legelinie wurde jeweils die Hälfte der Tiere in Bodenhaltung bzw. in Einzelkäfigen gehalten. Das Brustbein von allen Tieren wurde regelmäßig geröntgt. Die beiden braunlegenden Legelinien hatten mehr Brustbeinfrakturen als die drei weißlegenden Linien, während die Deformationen bei den Weißlegern tendenziell größer waren. Innerhalb der Braunleger zeigte die Hochleistungslinie mehr Frakturen und Deformationen als die Minderleistungslinie. Die Prävalenz von Frakturen war in der Bodenhaltung höher als in Einzelkäfigen, während innerhalb einiger Legelinien die Deformationen im Käfig größer waren als in der Bodenhaltung. Die Frakturprävalenz nahm mit dem Alter der Hennen zu. Die vorgestellte Röntgenmethode, die auch im Stall eingesetzt werden kann, erwies sich als eine zuverlässige und schnell durchzuführende Methode, um Brustbeinfrakturen und -deformationen wiederholt an denselben Tieren zu diagnostizieren und zu vermessen. Das Ziel der zweiten Studie war es, ein Tiermodell mit nicht-legenden Hennen zu entwickeln, welches die detaillierte Untersuchung des Einflusses der Legetätigkeit auf unterschiedliche Erkrankungen bei Hennen, u.a. Brustbeinschäden, erlaubt. Hierfür wurde jeweils zehn Hennen kurz nach sowie zehn Hennen vor Legebeginn ein subkutanes Implantat mit dem Gonadotropin-Releasing-Hormon (GnRH)-Agonisten Deslorelinazetat subkutan appliziert. Jeweils zehn weitere Hennen wurden als Kontrolltiere für beide Gruppen (nach / vor Legebeginn) gehalten. Das Implantat unterband die Legetätigkeit bei allen Hennen, die nach Legebeginn behandelt wurden, und zögerte den Legebeginn bei allen Hennen, die es davor erhalten hatten, hinaus. Die Wirkungsdauer war jedoch relativ kurz und es zeigte sich, dass die Gabe des Implantates im Abstand von ca. zwölf Wochen wiederholt werden sollte, um eine durchgängige Unterdrückung der Legetätigkeit zu erreichen. Des Weiteren zeigte sich ein niedrigerer 17β-Östradiol-Plasmaspiegel bei behandelten im Vergleich zu Kontrolltieren. Die Hennen dieses Versuches wurden ebenfalls zweimal während des Versuchszeitraums geröntgt. Innerhalb der Gruppe, die das Implantat nach Legebeginn erhielt, zeigten die legenden Hennen (Kontrolltiere) signifikant mehr Brustbeinfrakturen und größere Deformationen als die nicht-legenden Hennen. Die Größe der Deformationen nahm in dieser Gruppe insgesamt mit zunehmendem Alter der Hennen zu. Die Ergebnisse aus diesen beiden Studien wurden für die dritte Studie genutzt, deren Ziel es war, den Einfluss der Legetätigkeit sowie von 17β-Östradiol auf die Entstehung von Brustbeinschäden zu untersuchen. Es wurden jeweils 100 Hennen einer Hochleistungs- und 100 Hennen einer Minderleistungslinie in Bodenhaltung gehalten. Bei jeweils der Hälfte der Tiere beider Legelinien wurde die Legetätigkeit durch wiederholte Gabe eines Deslorelinazetat-Implantates unterbunden. Ein Teil dieser sowie der legenden Hennen bekam zusätzlich ein Implantat mit dem Steroidhormon 17β-Östradiol. Das Auftreten und die Schwere von Brusbeinfrakturen und -deformationen wurden durch wiederholte Röntgenuntersuchungen beurteilt. Des Weiteren wurde die Röntgendichte des Brustbeins ermittelt. Die Wahrscheinlichkeit, innerhalb des Versuchszeitraumes eine Fraktur zu erleiden, war bei den nicht-legenden Hennen sehr viel niedriger als bei den legenden Hennen, während kein Effekt der Legetätigkeit auf Brustbeindeformationen festgestellt werden konnte. Die Röntgendichte des Brustbeins war am Ende des Versuchszeitraums bei den nicht-legenden Hennen höher als bei den legenden. Die Gabe von exogenem Östradiol führte nur innerhalb der legenden Hennen zu einem moderat erhöhten Frakturrisiko und hatte keinen Effekt auf das Auftreten von Brustbeindeformationen sowie auf die Röntgendichte. Die hochleistende Legelinie wies eine höhere Wahrscheinlichkeit für Brustbeinfrakturen auf als die minderleistende Legelinie, während sich die beiden Linien in Bezug auf Deformationen nicht unterschieden. Zusammenfassend lässt sich sagen, dass in der vorliegenden Arbeit sowohl eine Methode zur Beurteilung der Brustbeingesundheit in Verlaufsstudien als auch ein Tiermodell zur Untersuchung des Zusammenhangs der Legetätigkeit sowie des frühen Legebeginns und verschiedenen Erkrankungen bei Legehennen erfolgreich etabliert und präsentiert wurden. Des Weiteren wurden unterschiedliche Risikofaktoren für das Auftreten von Brustbeinfrakturen einerseits und -deformationen andererseits gefunden. Dies zeigt, dass es sich hierbei um zwei voneinander unabhängige Symptome zu handeln scheint, weshalb eine klare Abgrenzung zwischen den beiden immens wichtig ist. Es wurde gezeigt, dass sowohl der untersuchte exogene Faktor als auch die untersuchten endogenen Faktoren einen Einfluss auf die Entstehung von Brustbeinschäden haben. Ein Teil der Frakturen scheint durch Kollisionen mit Einrichtungsgegenständen zu entstehen. Der enorme Unterschied zwischen legenden und nicht-legenden Hennen in Bezug auf die Wahrscheinlichkeit, eine Fraktur zu erleiden, zeigt jedoch sehr deutlich, dass eine durch die Legetätigkeit verursachte Schwäche des Brustbeins besteht, die diesen Knochen sehr anfällig für Frakturen macht. Der festgestellte Unterschied zwischen Hoch- und Minderleistungslinien untermauert diese Schlussfolgerung. Es erscheint daher notwendig, das Brustbein in weiteren Studien genauer zu untersuchen, um Unterschiede in der Knochenstruktur und -zusammensetzung zwischen legenden und nichtlegenden Hennen zu finden, welche die unterschiedliche Frakturanfälligkeit bedingen. Basierend auf diesen Ergebnissen könnten neue Ansatzpunkte zur Vermeidung dieses gravierenden Tierschutzproblems entwickelt werden

Table2_Japanese quails (Cortunix Japonica) show keel bone damage during the laying period—a radiography study.XLSX

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.</p

Table1_Japanese quails (Cortunix Japonica) show keel bone damage during the laying period—a radiography study.XLSX

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.</p

Table3_Japanese quails (Cortunix Japonica) show keel bone damage during the laying period—a radiography study.XLSX

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.</p

A fractured keel bone with callus formation.

<p>The arrow shows the fracture callus.</p

A fractured keel bone without callus formation.

<p>The arrow shows the fracture, a thin, black line.</p

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

<div><p>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 35^th, 51^st and 72^nd 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 72^nd week of age (p<0.05). In the 72^nd 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 51^st and 72^nd 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.</p></div

Proportion of deviated keel bone area (POD) (back-transformed LSM and upper and lower bounds of the 95% confidence interval) in %.

<p>POD was affected by the three-way interaction between housing system, layer line and age (p<0.01).</p