Genomic analyses of behavior traits in laying hen lines divergently selected for feather pecking

Feather pecking is a longstanding problem in commercial layer flocks. It often causes injured birds and even cannibalism. In the past, hens were beak trimmed to reduce feather pecking. Nevertheless, this procedure is already prohibited in some EU countries. Hence, a solution to this problem is urgently needed. The experimental populations analyzed in this thesis were formed by hens based on a White Leghorn layer strain which were divergently selected for high and low feather pecking since 1995. The first experimental population of this thesis was an F2 cross of about 900 hens which was established of the 10th generation of the pure selection lines. The second population consisted of about 500 hens of the 15th generation of these two lines. The aim of this thesis was to gain further knowledge of the genetic background of feather pecking and its relation to additional behavior traits and the gut microbiome. In chapter one, a novel model to detect extreme feather pecking hens was developed. Therefore, a mixture of two negative binomial distributions was fitted to feather pecking data of the F2 cross. With the estimated parameters, the trait posterior probability of a hen to belong to the extreme feather pecking subgroup (pEFP) was calculated. The fear tests tonic immobility and emerge box were conducted at juvenile and adult age of the hens to relate fearfulness to pEFP. After dichotomization, all traits were analyzed in a multivariate threshold model and subsequent genomewide association studies (GWAS) were performed. The fit revealed that extreme feather peckers made up a proportion of about one third of the hens. The new trait pEFP has a medium heritability of 0.35 and is positively correlated with the fear traits. Breeding for this new trait could be an option to reduce the proportion of extreme feather peckers. An index of fear related traits might serve as a proxy to breed indirectly against pEFP. In chapter two, the model to detect extreme feather pecking hens was applied to the pure selection lines. After calculation of the trait pEFP, GWAS with a subsequent post GWAS analysis were performed. Additionally, to find genomic regions influencing feather pecking, selection signatures were mapped by applying the intra-population iHS and the inter-population FST approach. Mapping of selection signatures revealed no clear regions under selection. GWAS revealed a region on chromosome one, where the existence of a quantitative trait locus (QTL) influencing feather pecking is likely. The candidate genes found in this region are a part of the GABAergic system. Despite the polygenic nature of feather pecking, selection on these candidate genes may reduce the extreme occurrence of it. In chapter three, the relation between agonistic behavior and feather pecking was analyzed. Therefore, the active parts of the traits (delivery of feather pecking, aggressive pecking or threatening) as well as the passive parts (reception of the traits) were considered. These groups of traits were additionally summarized by means of an index formation which led to the two additional traits Activity and Passivity, because all these behaviors are undesired in their excessive manifestations. Moreover, Indices were built by subtracting the passive traits from the respective active traits to obtain the feather pecking index, the aggression index and the threat index. Phenotypic correlations were estimated between all traits which were followed by heritability estimations and GWAS. Feather pecking is significantly positively correlated with the agonistic traits in both lines. The active traits and the feather pecking index show medium heritabilities. Hence, selection on high feather pecking leads to an increase of agonistic behavior whereas the correlation probably depends on the phase of establishing the social hierarchy and might disappear, after a stable ranking is established. GWAS revealed that the heritable traits in this study seem to be typical quantitative traits. Chapter four provides the analyses of the gut microbial composition of the two feather pecking lines, followed by the estimation of microbiabilities for feather pecking and the two agonistic behavior traits, to study the influence of the gut microbiome on behavior. Microbiota samples from digesta and mucosa were taken from ileum and caecum. The microbial communities were determined by using 16S RNA gene sequencing techniques. Although both lines differ significantly in some fractions of their gut microbial composition, the microbial animal effects were mostly negligibly small. Thus, the calculated microbiabilities were close to zero and not significant in both lines and for all traits investigated. Hence, trait variations were not affected by the gut microbial composition in both feather pecking lines. The thesis ends with a general discussion where additional results of a meta-analysis of pEFP and breeding strategies against feather pecking are considered.Federpicken ist ein lange bestehendes Problem in kommerziellen Legehennenherden. Es führt häufig zu Verletzungen und sogar Kannibalismus. In der Vergangenheit wurden Schnäbel gekürzt, um das Federpicken zu reduzieren. Da dieses Verfahren in einigen EU-Ländern bereits verboten ist, ist eine Lösung dringend erforderlich. Die analysierten Versuchspopulationen bildeten Hennen, die auf einer Weißen Leghorn Legerasse basierten und seit 1995 divergent für hohes und niedriges Federpicken selektiert wurden. Die erste Versuchspopulation war eine F2-Kreuzung von etwa 900 Hennen, die aus der 10. Generation der reinen Selektionslinien gebildet wurde. Die zweite Population bestand aus etwa 500 Hennen der 15. Generation dieser beiden Linien. Das Ziel war es, weitere Erkenntnisse über den genetischen Hintergrund des Federpickens und dessen Beziehung zu weiteren Verhaltensmerkmalen sowie dem Darmmikrobiom zu gewinnen. Im ersten Kapitel wurde ein neuartiges Modell zum Nachweis extremen Federpickens ausgearbeitet. Dazu wurde eine Mischung aus zwei negativen Binomialverteilungen an die Federpickdaten der F2-Kreuzung angepasst. Mit den geschätzten Parametern wurde das Merkmal die a posteriori Wahrscheinlichkeit einer Henne, zur Untergruppe der extremen Federpicker zu gehören (pEFP), berechnet. Die Furchttests tonische Immobilität und Emerge Box wurden in juvenilem und adultem Alter durchgeführt, um die Furcht mit pEFP in Beziehung zu setzen. Nach der Dichotomisierung wurden alle Merkmale in einem multivariaten Schwellenwertmodell analysiert und anschließend genomweite Assoziationsstudien (GWAS) durchgeführt. Extreme Federpicker machten etwa ein Drittel der Hennen aus. Das neue Merkmal pEFP hat eine mittlere Heritabilität von 0,35 und ist positiv mit den Furchtmerkmalen korreliert. Die Züchtung dieses neuen Merkmals könnte den Anteil extremer Federpicker reduzieren. Ein Index der furchtbezogenen Merkmale könnte als Hilfsmerkmal dienen, um indirekt gegen pEFP zu züchten. Im zweiten Kapitel wurde das Modell zum Nachweis extremer Federpicker auf die zwei reinen Selektionslinien angewandt. Nach der Berechnung des Merkmals pEFP wurden GWAS mit einer anschließenden post GWAS Analyse durchgeführt. Um zusätzlich genomische Regionen zu detektieren die das Federpicken beeinflussen, wurden Selektionssignaturen mittels intra- (iHS) und inter-Populations-Ansatz (FST) kartiert. Diese Kartierung ergab keine eindeutigen Regionen, an denen Selektion stattgefunden hat. Die GWAS zeigte eine Region auf Chromosom eins, in der die Existenz eines quantitative trait locus, welcher Federpicken beeinflusst, wahrscheinlich ist. Die gefundenen Kandidatengene sind ein Teil des GABA-Systems. Trotz der polygenen Natur des Merkmals Federpicken könnte die Selektion auf diese Kandidatengene das extreme Auftreten des Federpickens reduzieren. In Kapitel drei wurde die Beziehung zwischen agonistischem Verhalten und Federpicken analysiert. Dabei wurden die aktiven (Ausübung des Federpickens, aggressiven Pickens oder Drohens) und die passiven (Empfang der Merkmale) Anteile der Merkmale betrachtet. Diese Merkmalsgruppen wurden in einer Indexbildung zusammengefasst, die zu den beiden Merkmalen Aktivität und Passivität führte, da all diese Verhaltensweisen in ihrer exzessiven Ausprägung unerwünscht sind. Es wurden Indizes gebildet, indem die passiven von den jeweiligen aktiven Merkmalen subtrahiert wurden, um den Federpick-, Aggressions- und Bedrohungsindex zu erhalten. Zwischen allen Merkmalen wurden phänotypische Korrelationen und Heritabilitäten geschätzt und GWAS angewandt. Federpicken ist signifikant positiv mit den agonistischen Merkmalen in beiden Linien korreliert. Die aktiven Merkmale und der Federpick-Index zeigen mittlere Heritabilitäten. Daher führt die Selektion auf hohes Federpicken zu einer Zunahme des agonistischen Verhaltens, wobei die Korrelation wahrscheinlich von der Phase der Etablierung der sozialen Hierarchie abhängt. Die GWAS ergab, dass es sich um typische quantitative Merkmale zu handeln scheint. Kapitel vier enthält die Analysen der Darmmikrobiota der beiden Linien, sowie die Schätzung der Microbiabilities für Federpicken und agonistischer Verhaltensmerkmale, um den Einfluss des Darmmikrobioms auf das Verhalten zu untersuchen. Mikrobiotaproben aus der Digesta und Mucosa wurden aus Ileum und Caecum entnommen. Die Mikrobengemeinschaften wurden mit Hilfe von 16S-RNA Gen-Sequenzierungstechniken bestimmt. Obwohl sich beide Linien in einigen Fraktionen ihrer mikrobiellen Zusammensetzung im Darm signifikant unterscheiden, waren die mikrobiellen Tiereffekte meist vernachlässigbar gering. Somit waren die berechneten Microbiabilities nahe Null und nicht signifikant. Dies bedeutet, dass die Merkmalsvariation nicht durch die Zusammensetzung des Darmmikrobioms beeinflusst wurde. Die Dissertation endet mit einer allgemeinen Diskussion, in der zusätzliche Ergebnisse einer Meta-Analyse sowie Zuchtstrategien gegen Federpicken berücksichtigt werden

Meta-analyses of genome wide association studies in lines of laying hens divergently selected for feather pecking using imputed sequence level genotypes.

BACKGROUND: Feather pecking (FP) is damaging behavior in laying hens leading to global economic losses in the layer industry and massive impairments of animal welfare. The objective of the study was to discover genetic variants and affected genes that lead to FP behavior. To achieve that we imputed low-density genotypes from two different populations of layers divergently selected for FP to sequence level by performing whole genome sequencing on founder and half-sib individuals. In order to decipher the genetic structure of FP, genome wide association studies and meta-analyses of two resource populations were carried out by focusing on the traits 'feather pecks delivered' (FPD) and the 'posterior probability of a hen to belong to the extreme feather pecking subgroup' (pEFP). RESULTS: In this meta-analysis, we discovered numerous genes that are affected by polymorphisms significantly associated with the trait FPD. Among them SPATS2L, ZEB2, KCHN8, and MRPL13 which have been previously connected to psychiatric disorders with the latter two being responsive to nicotine treatment. Gene set enrichment analysis revealed that phosphatidylinositol signaling is affected by genes identified in the GWAS and that the Golgi apparatus as well as brain structure may be involved in the development of a FP phenotype. Further, we were able to validate a previously discovered QTL for the trait pEFP on GGA1, which contains variants affecting NIPA1, KIAA1211L, AFF3, and TSGA10. CONCLUSIONS: We provide evidence for the involvement of numerous genes in the propensity to exhibit FP behavior that could aid in the selection against this unwanted trait. Furthermore, we identified variants that are involved in phosphatidylinositol signaling, Golgi metabolism and cell structure and therefore propose changes in brain structure to be an influential factor in FP, as already described in human neuropsychiatric disorders

DIFFERENTIAL PHYSIOLOGICAL RESPONSES TO SOCIAL-EVALUATIVE STRESS IN DEPRESSIVE PATIENTS WITH VARIOUS DEGREES OF PEER VICTIMIZATION

Iffland B, Kley H, Neuner F. DIFFERENTIAL PHYSIOLOGICAL RESPONSES TO SOCIAL-EVALUATIVE STRESS IN DEPRESSIVE PATIENTS WITH VARIOUS DEGREES OF PEER VICTIMIZATION. Psychophysiology . 2023;60(Suppl. 1):S12

Distinct physiological responses to social-evaluative stress in patients with major depressive disorder reporting a history of peer victimization

Iffland B, Kley H, Neuner F. Distinct physiological responses to social-evaluative stress in patients with major depressive disorder reporting a history of peer victimization. Biological Psychology. 2023: 108697.Peer victimization is a risk factor for the development of major depressive disorders, but little is known about the mechanisms. This study examined whether peer victimization alters physiological and affective responses to potentially threatening social stimuli. For this purpose, reactions to socially evaluative stimuli of depressive patients and healthy controls with varying histories of peer victimization were compared. In a social conditioning task, we studied heart rate responses to unconditioned socially negative and neutral evaluative video statements, followed by the heart rate reactions to conditioned stimuli, i.e. still images of the faces of the same actors. Diagnosis of depression and peer victimization were both associated with a more pronounced heart rate deceleration in response to unconditioned stimuli, irrespective of valence. The effect of peer victimization was stronger in depressive patients than in healthy controls. However, heart rate responses to the CSs were not related to depression or peer victimization. The results indicate a hypervigilant processing of social stimuli in depressive patients reporting histories of peer victimization. This distinct processing may be associated with inappropriate behavioral and emotional responses to social challenges, putting individuals at risk for depressive symptoms. Copyright © 2023. Published by Elsevier B.V

Childhood Maltreatment in Individuals With Schizophrenia Spectrum Disorders: The Impact of Cut-Off Scores on Prevalence Rates

Weitkämper A, Kellner M, Iffland JR, et al. Childhood Maltreatment in Individuals With Schizophrenia Spectrum Disorders: The Impact of Cut-Off Scores on Prevalence Rates. Frontiers in Psychiatry. 2021;12:692492.Childhood maltreatment is a common phenomenon in various psychiatric disorders. Accordingly, patients with disorders from the schizophrenia spectrum (SSD) appear to have high prevalence rates of childhood maltreatment. However, the interpretation and comparability of prevalence rates is impeded by methodological weaknesses and differences such as measures and thresholds used in previous studies. Therefore, we aimed to provide and compare systematically captured data on prevalence rates of all common types of childhood maltreatment in patients with SSD using a standardized and well-established questionnaire and the most frequently used thresholds. The sample consisted of 48 patients with a primary diagnosis of SSD. 58.3–77.1% of the sample experienced at least one type of childhood maltreatment. Prevalence rates for physical abuse, physical neglect, and emotional abuse were dependent on the thresholds used, while equal rates were found for emotional neglect and sexual abuse. Physical neglect (46–67%), and emotional abuse (44–48%) were most commonly reported, followed by emotional neglect (38%), physical abuse (25–38%), and sexual abuse (25%). Additionally, high levels of peer victimization were reported by SSD patients. It appears that childhood maltreatment is a common phenomenon in SSD, even though methodological details, especially cut-off scores, have a substantial impact on the prevalence rates that are determined. Therefore, the methodology of studies should be closely examined when drawing conclusions from presented prevalence rates

Growing relevance of reports of adolescent cyberbullying victimization among adult outpatients

Iffland B, Bartsch LM, Kley H, Neuner F. Growing relevance of reports of adolescent cyberbullying victimization among adult outpatients. BMC Public Health. 2023;23(1): 1503.**Abstract** In the general population, prevalence rates of cyberbullying victimization have continuously increased over the past decades. However, the extent to which these increasing numbers affect clinical populations seeking treatment in outpatient services remains an open question. The present study sought to examine whether the increase of cyberbullying victimization is also reflected by increased reports of cyberbullying victimization in a clinical outpatient population. In addition, we assessed the incremental contribution of experiences of cyberbullying in the prediction of psychological symptoms when controlling for histories of childhood maltreatment and offline peer victimization. For this purpose, we analyzed routine data fromN = 827 outpatients who had sought treatment at a University outpatient clinic for psychotherapy between 2012 and 2021 in a cross-sectional study design. Analyses showed that 8.3% of the patients born in the years 1980 to 2002 indicated the experience of cyberbullying victimization in their adolescence. The rate of reported cyberbullying victimization increased from 1 to 3% in patients born in the years 1980 to 1987 to 24% in patients born in the year 2000. A logistic regression revealed that patients born in the years 1995–2002 were up to nineteen times as likely to report cyberbullying victimization as patients born in the years 1980–1982. In addition, hierarchical multiple regression analyses indicated that cyberbullying victimization significantly accounted for an incremental proportion of variance (1%) in the prediction of psychological symptom distress after controlling for child maltreatment and offline peer victimization. In conclusion, this retrospective survey indicates an increase of the clinical relevance of cyberbullying victimization both in frequency of and potential contribution to etiology. Raising attention to cyberbullying in clinical care and research seems to be justified and warranted

“She finds you abhorrent” - The impact of emotional context information on the cortical processing of neutral faces in depression

Iffland B, Klein F, Schindler S, Kley H, Neuner F. “She finds you abhorrent” - The impact of emotional context information on the cortical processing of neutral faces in depression. Cognitive, Affective, & Behavioral Neuroscience. 2021;21(2):426–444.**Abstract** Depression is associated with abnormalities in patterns of information processing, particularly in the context of processing of interpersonal information. The present study was designed to investigate the differences in depressive individuals in cortical processing of facial stimuli when neutral faces were presented in a context that involved information about emotional valence as well as self-reference. In 21 depressive patients and 20 healthy controls, event-related potentials (ERPs) were recorded during the presentation of neutral facial expressions, which were accompanied by affective context information that was either self- or other-related. Across conditions, depressive patients showed larger mean P100 amplitudes than healthy controls. Furthermore, mean late positive potential (LPP) amplitudes of depressive patients were larger in response to faces in self-related than in other-related context. In addition, irrespective of self-reference, mean LPP responses of depressive patients to faces presented after socially threatening sentences were larger compared with faces presented after neutral sentences. Results regarding self-reference supported results of previous studies indicating larger mean amplitudes in self-related conditions. Findings suggest a general heightened initial responsiveness to emotional cues and a sustained emotion processing of socially threatening information in depressive patients

Insights into the Mitochondrial and Nuclear Genome Diversity of Two High Yielding Strains of Laying Hens

Mitochondria are essential components of eukaryotes as they are involved in several organismic key processes such as energy production, apoptosis and cell growth. Despite their importance for the metabolism and physiology of all eukaryotic organisms, the impact of mitochondrial haplotype variation has only been studied for very few species. In this study we sequenced the mitochondrial genome of 180 individuals from two different strains of laying hens. The resulting haplotypes were combined with performance data such as body weight, feed intake and phosphorus utilization to assess their influence on the hens in five different life stages. After detecting a surprisingly low level of genetic diversity, we investigated the nuclear genetic background to estimate whether the low mitochondrial diversity is representative for the whole genetic background of the strains. Our results highlight the need for more in-depth investigation of the genetic compositions and mito-nuclear interaction in individuals to elucidate the basis of phenotypic performance differences. In addition, we raise the question of how the lack of mitochondrial variation developed, since the mitochondrial genome represents genetic information usually not considered in breeding approaches

Genetic Analyses and Genome-Wide Association Studies on Pathogen Resistance of <i>Bos taurus</i> and <i>Bos indicus</i> Cattle Breeds in Cameroon

Autochthonous taurine and later introduced zebu cattle from Cameroon differ considerably in their resistance to endemic pathogens with little to no reports of the underlying genetic make-up. Breed history and habitat variations are reported to contribute significantly to this diversity worldwide, presumably in Cameroon as well, where locations diverge in climate, pasture, and prevalence of infectious agents. In order to investigate the genetic background, the genotypes of 685 individuals of different Cameroonian breeds were analysed by using the BovineSNP50v3 BeadChip. The variance components including heritability were estimated and genome-wide association studies (GWAS) were performed. Phenotypes were obtained by parasitological screening and categorised in Tick-borne pathogens (TBP), gastrointestinal nematodes (GIN), and onchocercosis (ONC). Estimated heritabilities were low for GIN and TBP (0.079 (se = 0.084) and 0.109 (se = 0.103) respectively) and moderate for ONC (0.216 (se = 0.094)). Further than revealing the quantitative nature of the traits, GWAS identified putative trait-associated genomic regions on five chromosomes, including the chromosomes 11 and 18 for GIN, 20 and 24 for TBP, and 12 for ONC. The results imply that breeding for resistant animals in the cattle population from Northern Cameroon might be possible for the studied pathogens; however, further research in this field using larger datasets will be required to improve the resistance towards pathogen infections, propose candidate genes or to infer biological pathways, as well as the genetic structures of African multi-breed populations