Browsing by Person "Stefanski, Volker"

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Diurnal and photoperiodic effects on the immune system and glucocorticoid signaling in domestic pigs
(2019) Engert, Larissa; Stefanski, Volker
Physiology and behavior of humans and animals display pronounced diurnal and seasonal rhythmic variations. Diurnal rhythms are controlled by daylight and seasonal rhythms are adjusted by the photoperiod, i.e., the relative span of light per day. Modern human life and housing conditions of livestock are often directed against natural daylight conditions and thus, may entail circadian disruption causing misalignment between the central circadian pacemaker and peripheral tissues. Thereby, glucocorticoids are regarded as a main link between these compartments. Circadian disruption might be detrimental for health, in particular affecting immune function, which was mainly investigated in humans and nocturnal rodents but underlying mechanisms are not clearly defined yet. Therefore, the main objective of the present thesis was to investigate diurnal and photoperiodic effects on the immune system and glucocorticoid signaling as well as potential underlying endocrine, behavioral, and molecular mechanisms of these effects in domestic pigs. Adult male castrated pigs, held under specific lighting schedules, were surgically catheterized to enable blood collection without disturbance of the animals. Initially, domestic pigs were held under standard 12L:12D-lighting conditions and blood samples were taken every 2 hours over periods of up to 50 hours. Cosinor analyses revealed pronounced diurnal rhythmicity in peripheral leukocyte numbers of various immune cell populations. These rhythms were mainly comparable to results in humans and nocturnal rodents in relation to their respective rest-activity cycles, with the exception of porcine neutrophils differing from both species. Moreover, the investigated diurnal rhythms in activity behavior and plasma cortisol concentration confirmed that domestic pigs under the applied experimental conditions are diurnally active like humans. Linear mixed model analyses revealed associations of immune cell counts with plasma cortisol concentration, which also resembles results from humans and rodents. Subsequently, photoperiodic effects on diurnal rhythms in peripheral immune cell numbers were investigated for the first time in any species. Domestic pigs were held either under long day conditions (LD) or under short day conditions (SD) and were sampled every 2 hours over periods of 50 hours. Distinct photoperiodic differences in relative amplitudes and peak times of cell counts in various porcine leukocyte types were found, whereas mesor values did not differ. Moreover, photoperiodic effects on diurnal rhythms in plasma cortisol concentrations and activity behavior were found, which is in agreement with human and primate studies. Generalized linear mixed model analyses again revealed associations of leukocyte counts with plasma cortisol concentration and with activity behavior as well. In summary, the results imply stronger rhythmicity of peripheral immune cell numbers in general under SD than under LD. Common intrinsic mechanisms seem to regulate diurnal rhythms in peripheral leukocyte numbers in most immune cell types in domestic pigs, except for neutrophils again. Finally, to investigate potential molecular differences in diurnal regulation between different immune cell types, glucocorticoid receptor (GR) number and affinity were examined in peripheral blood mononuclear cells (PBMC) and granulocytes of domestic pigs. Thereby, a greater number of GR sites per cell and a higher GR binding affinity in PBMC compared to granulocytes were found, pointing to differences in the molecular mechanisms of glucocorticoid signaling between leukocyte populations. The results of the present thesis project are subsequently discussed in regard to specific implications for immune function and health as well as animal husbandry and welfare. Moreover, a methodological assessment of the approaches used within the thesis project was carried out and finally, suggestions for future research directions were given. In conclusion, the present thesis revealed for the first time diurnal and photoperiodic effects on the immune system as well as glucocorticoid signaling in domestic pigs and uncovered potential underlying mechanisms of these effects. Hence, an additional diurnally active model species in chronoimmunology research was established. Moreover, conducting chronoimmunology research in the porcine species represents an innovative approach in agricultural science and provides entirely new opportunities to improve animal health and welfare. Thereby, future studies might investigate diurnal differences in immune function, clarify the role of different zeitgebers on immune rhythms, and assess potential consequences of stressor exposure at different times of the day.
Does carrying a rider change motor and sensory laterality in horses?
(2022) Schwarz, Sophie; Marr, Isabell; Farmer, Kate; Graf, Katja; Stefanski, Volker; Krueger, Konstanze
Laterality in horses has been studied in recent decades. Although most horses are kept for riding purposes, there has been almost no research on how laterality may be affected by carrying a rider. In this study, 23 horses were tested for lateral preferences, both with and without a rider, in three different experiments. The rider gave minimal aids and rode on a long rein to allow the horse free choice. Firstly, motor laterality was assessed by observing forelimb preference when stepping over a pole. Secondly, sensory laterality was assessed by observing perceptual side preferences when the horse was confronted with (a) an unfamiliar person or (b) a novel object. After applying a generalised linear model, this preliminary study found that a rider increased the strength of motor laterality (p = 0.01) but did not affect sensory laterality (p = 0.8). This suggests that carrying a rider who is as passive as possible does not have an adverse effect on a horse’s stress levels and mental state.
Immunocastration of male pigs
(2020) Kress, Kevin Benjamin; Stefanski, Volker
In Europe, male piglets have been surgically castrated for centuries in order to avoid boar-tainted carcasses and to eliminate boar-specific aggressive and sexual behavior. Surgical castration of male piglets is still legal within the European Union during the first week of life, even without anaesthesia or analgesia. These circumstances have led to increasing societal criticism, as the castration is painful and violates the physical integrity of the animals. In 2010, European stakeholders of the pork chain committed themselves voluntarily to ending surgical castration from 2018 onwards, but at present, more than 2 years later, the majority of male piglets are still castrated surgically without adequate pain relief. Immunocastration is one alternative to surgical castration or pork production with boars. Although this technique is approved for commercial use in Europe since 2009, the market shares of immunocastrates within the European pork market are very low. The main reasons for this low market acceptance are uncertainties whether immunocastration is reliable, competitive and accepted along the pork chain. The main objective of the present doctoral thesis was therefore to investigate the advantages and disadvantages of immunocastration with regard to the three pillars of sustainability aspects. The various sustainability aspects of immunocastration were summarized and reviewed. In addition, the effects of different housing conditions on the reliability of immunocastration were experimentally tested, as was the impact of housing conditions and sex group on the growth performance of male pigs. Finally, the consequences of pork production with immunocastrates and boars on the profitability of German pig production were analyzed with a simulation model using different carcass pricing systems for immunocastrates and risk scenarios for boar taint. Immunocastration is an active immunization against GnRH and consists of two consecutive vaccinations to induce antibodies which temporarily suppress testicular functions and prevent boar taint. It is a method which ensures both a high product quality and a high level of animal welfare. The impact of immunocastration on the three pillars of sustainability has been studied extensively, although a contemporary global overview of its different aspects has been missing. Performance results in immunocastrates are better than in barrows, but worse than in boars. The environmental impact of pork production with immunocastrates is lower than with barrows, but higher than with boars. The level of aggression is considerably lower in immunocastrates compared to boars. Societal concerns are mainly related to food safety, and are not supported by scientific evidence. After the second vaccination, immunocastrates switch physiologically from a boar-like to a barrow-like status. Therefore, the timing of the second vaccination is a fine-tuning tool to balance advantages of boars with their environmental and economic benefits against the increased risk of welfare problems and boar taint. Nevertheless, both the synergic and the conflicting relationships between the pillars of sustainability must be communicated along the value chain to produce tailored pork products. The present doctoral thesis shows that immunocastration can balance the advantages and disadvantages of pork production with boars or barrows, and thus serve as a sustainable alternative for the European pork chain. If used correctly, immunocastration is reliable in preventing boar taint and can be economically competitive with traditional surgical castration. Based on this thesis, future studies might investigate quality assurance systems that reliably detect non-responders, or animals that are incorrectly vaccinated, before slaughter or at slaughter line. In addition, the economic impact of switching from traditional pork production with barrows to pork production with immunocastrates along the entire pork chain should be further analyzed.
Impact of dietary phosphorus and fermentable substrates on the immune system and the intestinal microbiota of the pig
(2016) Heyer, Charlotte Maria Elisabeth; Stefanski, Volker
Phosphorus (P) represents a crucial input for agriculture and food industries as a mineral present in ingredients used for livestock feeding as well as in mineral fertilisers. In the current systems, P is primarily derived from the finite mined phosphate rock resource. Thus, a critical challenge of global P scarcity is directly linked to future food security and sustainable resource management, especially in the European Union which is dependent on raw P from outside Europe. Apart from other future activities in animal nutrition, new dietary formulations of livestock diets emerged as a potential approach to increase the digestibility of plant P, phytate (myo inositol 1,2,3,4,5,6 hexakisphosphate, InsP6), and to reduce the supplementation with mineral phosphate. In non-ruminant animals, such as the pig, InsP6 hydrolysis is incomplete, as the small intestine lacks sufficient enzymes such as endogenous mucosal phytase and phosphatase. As a consequence, there is rising scientific interest to improve the understanding of InsP6 degradation in the digestive tract as well as the effects on nutritional factors and finally animal performance and health. The aim of the present thesis was to investigate the impact of dietary P, InsP6 and InsP6 hydrolysis products in combination with different fermentable substances (protein, carbohydrate) on the porcine immune system, the intestinal microbiota and animal health. First, a comprehensive literature overview describes the impact of P on the immune system and the microbiota along the gastrointestinal tract (GIT), including potential effects on host health with special focus on the pig. Secondly, an in vivo study with growing pigs was conducted to examine the effects of diets with varying mineral calcium-phosphorus (CaP) levels as well as different fermentable substrates on intestinal CaP concentration, InsP6 hydrolysis, the intestinal microbial ecosystem, and the peripheral and gut-associated immune system. In 2 consecutive experiments, 31 growing pigs (55 ± 4 kg) were allotted to a 2 × 2 factorial arrangement with 4 treatment groups, fed either a corn-soybean meal or a corn-pea based diet, each with 2 different CaP levels (low, 66% of the CaP requirement; high, 120% of the CaP requirement) supplemented with monocalcium phosphate and calcium carbonate. After 3 weeks of adaptation to the diets, all pigs were immunized twice with keyhole limpet hemocyanin (KLH). Blood and faeces samples were taken. After slaughtering, immunological tissue (jejunal, ileal mesenteric lymph nodes, spleen) as well as jejunal, ileal, caecal and colonic digesta were taken. Faecal and digesta samples were examined for P, Ca, inositol phosphate (InsP) isomers and for the marker titanium dioxide. The number of different leukocyte subpopulations analysed by flow cytometry, mitogen-induced lymphocytes proliferation in vitro were assessed. In addition, concentrations of plasma anti KLH IgM and plasma anti-KLH IgG analysed by ELISA and haematological parameters analysed by an automated hematology system have been measured in blood and tissue samples. In digesta samples, bacterial 16S rRNA gene copy numbers were determined by quantitative real-time PCR. The concentration of short chain fatty acids (SCFA) and ammonia was assessed. In addition, the use of terminal restriction fragment length polymorphism has been proven to characterize the structure of porcine gut microbiota. Results of the current study demonstrated that CaP and fermentable substrates had a distinct effect on the peripheral and gut-associated immune system, as well as on microbial composition and activity in growing pigs. High dietary CaP concentrations and the corn-pea diets increased P net absorption. Almost no InsP6 degradation could be observed in the GIT, and mainly myo inositol pentakisphosphate (InsP5) isomers were measured in jejunal, caecal digesta and faecal samples. In particular, the high CaP diets showed higher InsP6 and InsP5 concentrations, indicating a reduction of the initial steps of P release from InsP6 and a further breakdown of InsP5 isomers. The low CaP content might cause an impaired first line of defence and activation of the cellular and humoral adaptive immune response. As an example, the high CaP content affected the outcome of the adaptive immune response including a higher number of antigen experienced T-helper cells in the blood as well as higher plasma anti-KLH IgG concentrations. The reactivity of blood and mesenteric lymph node lymphocytes to Concanavalin A in these pigs was impaired, indicating modulating effects of other origin such as migration patterns or activity of antigen-presenting cells. Since results of the present study suggest contradictory effects of CaP level on immune cell numbers and lymphocyte reactivity in vitro and in vivo, further studies are needed to determine effects on cell signalling such as cytokine production profiles. Moreover, the high CaP content and the soybean meal diets increased the number of butyrate-producing bacteria, such as Eubacterium rectale and Roseburia spp. and increased the concentration of various SCFA in the small and large intestine, thereby contributing to improve gut health. Potentially harmful bacteria, such as Enterobacteriaceae and Bacteroides Prevotella Porphyromonas, were increased by the low CaP level and pea diets, indicating a less healthy microbiota. Results demonstrated that both, CaP supply and the amount of fermentable substrates, may beneficially affect gut health due to modulations of the composition and activity of the intestinal microbiota. Further studies should evaluate the impact of CaP on specific pathogenic bacteria known to produce toxic products creating a direct link to the immune system and animal health. Although most parameters of the present study indicate a positive effect of the high CaP diet, not all values showed a consistent effect on animal health, such as immune cell numbers and lymphocyte proliferation in vitro. In conclusion, variations in P availability and the formation of individual InsPs have to be considered when formulating diets in support of a stable intestinal microbial ecosystem and immune functions of the host.
Impact of housing environment and rearing condition on the immune system and welfare of laying hens
(2021) Hofmann, Tanja Melanie; Stefanski, Volker
In recent years, consumers’ interest in modern farm animal husbandry and, above all, criticism of intensive production systems that can impair animal welfare, have increased sharply. From hatch, chickens are confronted by a wide range of environmental and social stressors that may confound homeostasis and impair their health and welfare. Not being able to deal with the environment can lead to an activation of the stress system, resulting in a release of neuroendocrine signals like glucocorticoids. These have the potential to modify the immune system and alter species-specific behavior with possible detrimental impacts on animals’ health and welfare. In this context, early-life conditions play a special role as they were shown to have long-term consequences for physiology and behavior in the later life. High stocking densities which activate the hypothalamic-pituitary-adrenal-axis are said to be one of the largest welfare concerns in the poultry industry. Previous research has primarily focused on the effects of stocking density on broilers and adult laying hens, but few studies have focused explicitly on layer pullets, and none of them investigated long-term effects on the immune system. The understanding of short- and long-term consequences of environmental conditions during rearing on the immune system and behavior of layer pullets is essential for shaping an environment that avoids allostatic overload, thus promoting animal health and welfare. This doctoral thesis aimed to identify and evaluate potential immunomodulating factors in the housing environment of chickens. In order to achieve this, it summarizes and reviews important aspects of housing environments for the immune system in chickens. Furthermore, an existing flow cytometric method for the enumeration of leukocytes and discrimination of lymphocyte subsets in blood was adapted to lymphatic tissues. Moreover, short- and longterm consequences of stocking density during rearing on the immune system, stress hormone level and behavior in layer pullets were evaluated. The results are described in detail in three peer-reviewed manuscripts published in international journals. The literature analysis identified several housing conditions as immunomodulating factors. Light management was marked as an effective tool to modulate the immune response in chickens, as long light phases were demonstrated to lower adaptive cellular and humoral immune responses while certain light colors, especially of shorter wavelengths, promote some immune functions. Moreover, particularly high ammonia and hydrogen sulfide concentrations pose a threat to chickens’ health by dampening the adaptive immune response and promoting inflammation. However, the interaction between housing environment and management is complex. Depending on the type of housing form, chickens are confronted with different social and environmental challenges, which in turn can have an impact on the immune and stress system. An understanding of immunosuppressive risk factors is essential for successful poultry management aiming to optimize health, welfare and economic efficiency. To broaden the assessment of immune status for scientific purposes, new staining- and gating strategies for avian lymphatic tissues were adapted to an existing flow cytometric method for blood. The adaptation process showed that the application of already established protocols to other tissues is possible, but has to be examined carefully as tissue-specific effects or cell-preparative procedures might lead to differences in possible combinations of antibody-fluorochrome conjugates. The results of the experimental study within this doctoral thesis demonstrate for the first time that stocking density during rearing has not only short- but also long-term impacts on the immune system in laying hens. Pullets reared at high stocking densities showed stress-related alterations in immune cell numbers and higher incidences of feather pecking that manifest in the laying period. This highlights the significance of early-life conditions for immunocompetence throughout the whole production cycle. Of particular interest is that the number of gamma-delta T cells was lower in the high-density group, which has not been investigated in any stress-related studies with chickens before. The present doctoral thesis provides a significant contribution to the improvement of health and welfare in poultry farming and forms a basis for further research about long-lasting consequences of rearing conditions on the immune system, especially in laying hens.
Influence of different plant extracts on CYP-mediated skatole and indole degradation in pigs
(2024) Marro, Philipp; Wesoly, Raffael; Stefanski, Volker
One of the primary substances responsible for the unpleasant odor in boar meat is skatole. Enzymes belonging to the cytochrome P450 (CYP) family play a pivotal role in the hepatic clearance of skatole. This study aimed to investigate the impact of oregano essential oil (OEO), Schisandra chinensis extract (SC), and garlic essential oil (GEO) on hepatic CYP2E1 and CYP2A activity in pigs. In three consecutive trials, cannulated castrated male pigs were provided with a diet containing 0.2–0.3% of one of these plant extracts. Following a 14-day feeding period, the animals were slaughtered, and liver and fat samples were collected. The findings indicate that the activities of CYP2E1 were unaffected by any treatment. However, GEO treatment demonstrated a significant reduction in CYP2A activity (p < 0.05). Pigs treated with GEO also exhibited a notable increase in skatole concentrations in both plasma and adipose tissue. In contrast, animals fed SC displayed elevated skatole concentrations in plasma but not in fat tissue. OEO did not influence skatole concentrations in either blood or fat. Furthermore, the study revealed that a supplementation of 6 g GEO per animal per day induced a significant increase in skatole concentrations in blood plasma within 24 h.
Influence of frequent regrouping and social status on behavioral, endocrine and immune responses of group housed pregnant sows
(2020) Schalk, Christiane; Stefanski, Volker
In modern animal husbandry, dynamic group-housing of pregnant sows is a common practice. Every regrouping of animals or every change of group composition is associated with the establishment or the adjustment of a new dominance hierarchy, which provokes aggressive behavior, fights and injuries. This process is known to result in social stress by an activation of different stress systems. The subsequent release of neuroendocrine signals like glucocorticoids (e.g. cortisol) has the potential to alter several immune functions and immune cell numbers in the blood which may be directly associated with animals’ health, reproduction, embryonic development and economic losses. The effects of frequent regrouping or mixing on pregnant sows’ behavior, stress hormones and especially the distribution of blood leukocyte subpopulations represent a major research gap in the field of stress assessment of dynamic group-housing conditions in pig production. The aim of the present doctoral thesis was to evaluate whether frequent regrouping acts as a chronic social stressor influencing behavior as well as the endocrine and immune system of group-housed pregnant sows. Special emphasis was put on the question whether frequent changes of the group composition affect blood leukocyte subpopulations. A study with 40 pregnant sows was designed to investigate the influence of frequent changes of group composition on numbers of blood leukocyte subpopulations in combination with analyses of agonistic behavior and the endocrine status. Pregnant multiparous sows were housed in groups of five animals. Sows were either assigned to a repeated social mixing treatment with a mutual exchange of two randomly selected sows of two specific groups (2x2) twice a week over a period of eight weeks, or remained undisturbed in their original group. Blood samples of all sows were collected during pregnancy at five time points before, during, and after the mixing period to evaluate the number of blood leukocyte subpopulations and plasma cortisol concentrations. Blood immune cell numbers were analyzed during all trimesters of gestation and the impact of social status on these modifications was assessed. Behavioral data of pregnant sows of this experiment were used to compare various recommended dominance indices to rank individuals based on different methodical aspects to investigate whether these indices are comparable and equally applicable for determination of dominance relationships. Results of the current study demonstrated that pregnancy-associated alterations in the immune system generally exist in sows. The numbers of T cells, natural killer cells, B cells, cytotoxic T cells, and CD8+ γδ- T cells decreased during the last trimester of pregnancy, while neutrophils and plasma cortisol concentrations increased during pregnancy. Those pregnancy-associated alterations in the immune system were affected especially in middle-ranking sows, which had higher numbers of B cells and monocytes than sows with lower ranking positions. Plasma cortisol concentrations also tended to be higher in middle-ranking sows compared to low-ranking sows indicating that social rank can influence the immune system and endocrine status in sows during pregnancy. These findings showed the necessity to choose the appropriate measurement for calculation of dominance relationships. Repeated social mixing by frequent changes of group composition not only resulted in an increase of aggressive behavior during the entire mixing period, but also in altered immune cell numbers. The immunological profile in blood of mixed sows was characterized by lower numbers of antigen-experienced T helper cells, cytotoxic T cells and natural killer cells. This work demonstrated that frequent changes of group composition affect both cell numbers of the innate and the adaptive part of the immune system, which may weaken immunological memory functioning and reduce the resistance against certain infections in pregnant sows. For most of these immune cells a certain period of instable housing conditions was required to induce a change, but once manifested, these immunological alterations persisted even after the end of the mixing period. Although the findings of the present work on blood immune cell numbers resemble in many aspects a picture of stress-induced immunomodulation previously reported in context with social stress, no clear differences in measured plasma stress hormone concentrations between treatment groups or rank-positions were found. Whether other factors have influenced cortisol concentrations needs to be further evaluated. The overall picture emerging from the current doctoral thesis indicates that frequent changes of group composition and social status have the potential to induce stress-related immunological changes in pregnant sows which might adversely affect sows’ health.
Male reproductive organ weight: Criteria for detection of androstenone-positive carcasses in immunocastrated and entire male pigs
(2023) Fazarinc, Gregor; Batorek-Lukač, Nina; Škrlep, Martin; Poklukar, Klavdija; Van den Broeke, Alice; Kress, Kevin; Labussière, Etienne; Stefanski, Volker; Vrecl, Milka; Čandek-Potokar, Marjeta
Immunocastration and rearing of entire males (EMs) are sustainable alternatives to surgical castration. However, these animal carcasses have variable risk of boar taint and should be identified at the slaughter line. We aimed to identify a simple and reliable indicator of androstenone-related boar taint by evaluating pelvic urogenital tract weight as a marker of boar-taint animals at the slaughter line. The pelvic urogenital tract, testes, and accessory sex glands of EMs and immunocastrates (ICs) were collected, dissected, and weighed, before colorimetric measurements of testicular tissue. Additionally, GnRH antibody titers and testosterone, androstenone, and skatole levels were determined. Our results showed that 81.8% of EMs had androstenone levels above the risk threshold (>0.5 µg/g fat; EM/Ahigh subgroup), whereas in ICs, the C/Ahigh subgroup with androstenone >0.5 µg/g fat accounted for only 4.3%. Androstenone levels correlated negatively with GnRH antibody titers and positively with testosterone levels and reproductive organ weights. Identification of ICs with androstenone levels above the threshold (IC/Ahigh subgroup) may be achieved via testes or pelvic urogenital tract weight measurements. However, in EMs, the latter is a more reliable parameter. A principal component analysis based on these variables and hierarchical clustering also distinguished the Ahigh from the Alow subgroup, irrespective of IC/EM. The findings highlight the possible use of pelvic urogenital tract weight along with testes weight as a simple, reliable, and efficient morphometric indicator for identifying androstenone-positive carcasses of different sex categories.
Multi-omics reveals different strategies in the immune and metabolic systems of high-yielding strains of laying hens
(2022) Iqbal, Muhammad Arsalan; Reyer, Henry; Oster, Michael; Hadlich, Frieder; Trakooljul, Nares; Perdomo-Sabogal, Alvaro; Schmucker, Sonja; Stefanski, Volker; Roth, Christoph; Camarinha Silva, Amélia; Huber, Korinna; Sommerfeld, Vera; Rodehutscord, Markus; Wimmers, Klaus; Ponsuksili, Siriluck
Lohmann Brown (LB) and Lohmann Selected Leghorn (LSL) are two commercially important laying hen strains due to their high egg production and excellent commercial suitability. The present study integrated multiple data sets along the genotype-phenotype map to better understand how the genetic background of the two strains influences their molecular pathways. In total, 71 individuals were analyzed (LB, n = 36; LSL, n = 35). Data sets include gut miRNA and mRNA transcriptome data, microbiota composition, immune cells, inositol phosphate metabolites, minerals, and hormones from different organs of the two hen strains. All complex data sets were pre-processed, normalized, and compatible with the mixOmics platform. The most discriminant features between two laying strains included 20 miRNAs, 20 mRNAs, 16 immune cells, 10 microbes, 11 phenotypic traits, and 16 metabolites. The expression of specific miRNAs and the abundance of immune cell types were related to the enrichment of immune pathways in the LSL strain. In contrast, more microbial taxa specific to the LB strain were identified, and the abundance of certain microbes strongly correlated with host gut transcripts enriched in immunological and metabolic pathways. Our findings indicate that both strains employ distinct inherent strategies to acquire and maintain their immune and metabolic systems under high-performance conditions. In addition, the study provides a new perspective on a view of the functional biodiversity that emerges during strain selection and contributes to the understanding of the role of host–gut interaction, including immune phenotype, microbiota, gut transcriptome, and metabolome.
Non-invasive welfare evaluations in horses : the usefulness of laterality
(2020) Marr, Isabell; Stefanski, Volker
Animal welfare is becoming increasingly important, especially for animal owners. In particular, prey animals such as horses generally suffer in silence. To improve the evaluation of animal welfare in practice and to simplify it in scientific research, new welfare indicators are needed that are easier to assess, less time consuming, and repeatable. Therefore, this thesis aimed to investigate laterality as a welfare indicator that could meet these requirements. Domestic horses were used as model organisms as they display sensory and motor laterality on individual and/or population level; their sensory organs are placed laterally, non-invasive stress hormone analysis is already well-established, as a highly social animal that evolved on open ranges it is prone to suffering from inappropriate human management regimes, and it displays various stress responses. In STUDY it could be demonstrated that a shift to an enhanced preference for the right brain hemisphere was in line with increased stress hormone concentrations in faeces. Sensory laterality and motor laterality measured while grazing, shifted to the left when natural needs were restricted by a change from group to individual housing. Sensory laterality changed immediately after the change of housing conditions, whereas the motor laterality changed with a time delay of one week. STUDY 2 demonstrated that motor laterality measured as initial forelimb use correlated with the cognitive bias (welfare indicator). Right-sided horses were faster to approach an ambiguous stimulus and therefore displayed a positive cognitive bias. But neither motor laterality, measured through grazing stance, nor sensory laterality were related to cognitive bias. STUDY 3 demonstrated that a preference for left side sensory organ use is not only evident in negative contexts but also in positive contexts, because the horses also preferred their left side during affiliative interactions that are assumed to induce positive emotions. This study demonstrated that not only the direction of shift in laterality, but also the context of the shift, should be recorded to reliably identify poor or good welfare. Therefore, it is recommended that additional stress parameters be applied to reliably evaluate animal welfare. STUDY 4 investigated whether the sampling and analysis of faecal stress hormones and immunoglobulin A could be simplified by applying a novel conservation method. Often it is not possible to immediately freeze the faecal samples and/or the transportation to the lab is lengthy. The study demonstrated that faecal samples can be dried a closed system such as an air-tight tube containing silica gel. The samples were dried within 24 hours, as fast as in controlled air-drying conditions at room temperature. The new and simpler drying method prevented the stress hormones (glucocorticoid metabolites) from enzymatic degradation and conserved them, demonstrated by the fact that the detectable concentration remained unchanged. In contrast, immunoglobulin A showed a reduction in the detectable concentration. Therefore, if possible, the conservation of faecal samples should be avoided when immunoglobulin A is to be analysed, although it would be possible to apply an extrapolation to attain fairly reliable results. This new drying method will simplify research on wild horses into the type of stressors they are confronted with, the impact of natural stressors, and effect of stressors on, for example, their laterality in comparison with domestic horses. STUDY 5 investigated whether the strength of laterality provides insight into basal physiological and immunological status, stress response, stress reactivity, or cognitive bias. Only a correlation between age and the strength of laterality was found, with strength of laterality increasing with age. However, age could explain only 30 per cent of the inter-individual variation in strength of laterality. The results demonstrated that the strength of laterality is not a reliable indicator of animal welfare. The direction of laterality may be of greater importance. Altogether, it was demonstrated that laterality is a promising, reliable, repeatable, and objective indictor of animal welfare, which is quick and easy to asses, and inexpensive. Like other well-established welfare and stress indicators, laterality has its limitations. Therefore, it is recommended that other welfare indicator should be simultaneously assessed and changes in laterality recorded, as different traits and personalities result in a high inter-individual variation in base laterality indices. Possible influences and correlations between emotional processing and cerebral lateralization are discussed. Nonetheless, further research is needed to establish a more reliable measurement of motor laterality, and to better understand the relationship between emotional processing and lateralization, as well as possible influencing factors.
Stress hormone-induced immunomodulation and interplay between immune cells and bacteria in response to stress hormones in domestic pigs
(2020) Reiske, Lena; Stefanski, Volker
The two main endocrine systems involved in the regulation of stress reactions are the HPA axis, leading to the synthesis of glucocorticoids like cortisol or corticosterone, and the SAM axis, whose activation is associated with the release of the catecholamines adrenaline and noradrenaline. These stress hormones modulate the function of the immune system. Although pigs in modern husbandry systems face many stressors, the consequences of elevated plasma stress hormone levels on porcine immune cell numbers and functionality are insufficiently resolved. While some research on glucocorticoid effects has been conducted, data on many parameters are missing and catecholamines have not been studied systematically in the pig, yet. It is known that stress can negatively affect pigs’ resistance to infections like salmonellosis, but the underlying mechanisms are still subject to intense research efforts, with new perspectives arising since the discovery of interkingdom-signalling and microbial catecholamine perception. The aim of this thesis was to determine the effects of cortisol, adrenaline and noradrenaline on porcine immune cell functionality and the blood numbers of different leukocyte subsets. Furthermore, the interplay of immune cells and Salmonella Typhimurium under the influence of catecholamines was investigated. Adult male castrated pigs were surgically equipped with indwelling catheters to enable stress-free blood collection and intravenous application of hormones. In an initial experiment, the effects of in vitro stress hormone treatment on lymphocyte proliferation and the production of the proinflammatory cytokine TNFa were described. Cortisol reduced both proliferation and number of TNFa producers. Both catecholamines caused an increased lymphocyte proliferation at low concentrations whereas noradrenaline drastically decreased proliferation at high concentrations. While noradrenaline had no impact on TNFa producers, they were reduced in gd T cells and monocytes upon adrenaline addition. Overall, the effects were comparable to humans in terms of direction and dose but there were some disparities regarding adrenaline. In the second part of the project, the impact of in vivo stress hormone administration on immune cell numbers and functionality was examined by infusion for 48h. Cortisol and noradrenaline led to a decreased lymphocyte proliferation but to a variable extent and all three hormones promoted phagocytic function of innate immune cells. Cortisol caused a marked increase of neutrophil numbers while almost all other cell types declined strongly. For most cell types, noradrenaline exerted similar effects but solely after 2h whereas cortisol-induced alterations lasted the whole treatment period. Adrenaline effects were mostly reduced to CD8- T cells, which were reduced at first but increased after 24h. A sharp peak in NK cell numbers after 2h adrenaline infusion is particularly noteworthy and resembles findings from rodent and human studies. Overall, both hormone groups led to a shift from adaptive to innate immunity, underpinning the picture of a promotion of fast and unspecific defence systems to respond to threats in stressful situations. In a third study, S. Typhimurium was grown in the presence of catecholamines to determine the effects of supernatants from these cultures on porcine immune cell function. Both lymphocyte proliferation and TNFα production were hampered substantially, as opposed to the findings on catecholamine effects in the first experiment. It was demonstrated that these effects were not caused by catecholamines or their oxidation products and the formation of a so-far unknown immunosuppressive substance by catecholamine-primed bacteria was assumed. The results contribute to a better understanding of the increased susceptibility to infection in stressed animals and reveal a new dimension of cross-species communication. Finally, the results of the present thesis were discussed regarding their comparability to studies in humans and rodents and previous stress experiments in pigs. Furthermore, the effects of acute and chronic stress as well as different coping styles that are characterised by a SAM or HPA predominance on animal welfare and pig health were discussed, based on the endocrine mechanisms investigated in the present thesis. Possible implications of enhanced glucocorticoid and catecholamine levels for practical pig husbandry were given. Lastly, suggestions for future research to further elucidate the impact of stress hormones on the porcine immune system and the interplay with pathogenic bacteria were made.