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Palm, A. - K. E., Wattle, O., Lundström, T., & Wattrang, E. (2016). Secretory immunoglobulin A and immunoglobulin G in horse saliva. Vet. Immunol. Immunolpathol., 180, 59–65.
Abstract: This study aimed to increase the knowledge on salivary antibodies in the horse since these constitute an important part of the immune defence of the oral cavity. For that purpose assays to detect horse immunoglobulin A (IgA) including secretory IgA (SIgA) were set up and the molecular weights of different components of the horse IgA system were estimated. Moreover, samples from 51 clinically healthy horses were tested for total SIgA and IgG amounts in saliva and relative IgG3/5 (IgG(T)) and IgG4/7 (IgGb) content were tested in serum and saliva. Results showed a mean concentration of 74μg SIgA/ml horse saliva and that there was a large inter-individual variation in salivary SIgA concentration. For total IgG the mean concentration was approx. 5 times lower than that of SIgA, i.e. 20μg IgG/ml saliva and the inter-individual variation was lower than that observed for SIgA. The saliva-serum ratio for IgG isotypes IgG3/5 and IgG4/7 was also assessed in the sampled horses and this analysis showed that the saliva-serum ratio of IgG4/7 was in general approximately 4 times higher than that of IgG3/5. The large inter-individual variation in salivary SIgA levels observed for the normal healthy horses in the present study emphasises the need for a large number of observations when studying this parameter especially in a clinical setting. Moreover, our results also indicated that some of the salivary IgG does not originate from serum but may be produced locally. Thus, these results provide novel insight, and a base for further research, into salivary antibody responses of horses.
Keywords: Equine; Secretory IgA; IgG; Saliva; Mucosal immunity
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Maury, M., Murphy, K., Kumar, S., Mauerer, A., & Lee, G. (2005). Spray-drying of proteins: effects of sorbitol and trehalose on aggregation and FT-IR amide I spectrum of an immunoglobulin G. Eur. J. Pharm. Biopharm., 59(2), 251–261.
Abstract: An immunoglobulin G (IgG) was spray-dried on a Büchi 190 laboratory spray-dryer at inlet and outlet air temperatures of 130 and 190°C, respectively. The IgG solution contains initially 115mg/ml IgG plus 50mg/ml sorbitol. After dialysis, at least 80% of low molecular weight component was removed. After spray-drying the dialyzed IgG and immediate redissolution of the powder, an increase in aggregates from 1 to 17% occurred. A major shift towards increase β-sheet structure was detected in the spray-dried solid, which, however, reverted to native structure on redissolution of the powder. A correlation between aggregation determined by size exclusion chromatography and alterations in secondary structure determined by Fourier transformation infra-red spectroscopy could not therefore be established. On spray-drying a non-dialyzed, sorbitol-containing IgG only some 0.7% aggregates were formed. The sorbitol is therefore evidently able to stabilize partially the IgG during the process of spray-drying. Addition of trehalose to the liquid feed produced quantitatively the same stabilizing action on the IgG during spray-drying as did the sorbitol. This finding again points towards a water replacement stabilization mechanism. The IgG spray-dried powder prepared from the dialyzed liquid feed showed continued substantial aggregation on dry storage at 25°C. This was substantially less in the non-dialyzed, sorbitol-containing spray-dried powder. Addition of trehalose to both dialyzed and non-dialyzed system produced substantial improvement in storage stability and reduction in aggregate formation in storage. The quantitative stabilizing effect of the trehalose was only slightly higher than that of the sorbitol. Taken together, these results indicate that both the sorbitol and trehalose stabilize the IgG primarily by a water replacement mechanism rather than by glassy immobilization. The relevance of this work is its questioning of the importance of the usually considered dominance of glassy stabilization of protein in dried systems of high glass transition temperature, such as trehalose. The low glass transition temperature sorbitol produces almost equal process and storage stability in this case.
Keywords: Immunoglobulin; Spray-drying; Stabilization; Sorbitol; Trehalose; Water replacement
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Pawluski, J., Jego, P., Henry, S., Bruchet, A., Palme, R., Coste, C., et al. (2017). Low plasma cortisol and fecal cortisol metabolite measures as indicators of compromised welfare in domestic horses (Equus caballus). Plos One, 12(9), e0182257.
Abstract: The hypothalamic-pituitary-adrenal (HPA) axis response to chronic stress is far from straight forward, particularly with regards to animal welfare. There are reports of no effect as well as both decreases and increases in cortisol after chronic stressors. Therefore, the first aim of the present study was to determine how measures of compromised welfare, such as chronic pain and haematological anomalies, related to cortisol levels in domestic horses (Equus caballus). Domestic horses are an informative model to investigate the impact of chronic stress (due to environment, pain, work, housing conditions...) on the HPA axis. The second aim was to determine whether levels of fecal cortisol metabolites (FCM) may be used as an indicator of welfare measures. The present study used fifty-nine horses (44 geldings and 15 mares), from three riding centres in Brittany, France. The primary findings show that horses whose welfare was clearly compromised (as indicated by an unusual ears backward position, presence of vertebral problems or haematological anomalies, e.g. anaemia) also had lower levels of both FCM and plasma cortisol. This work extends our previous findings showing that withdrawn postures, indicators of depressive-like behavior in horses, are associated with lower plasma cortisol levels. We also found that evening plasma cortisol levels positively correlated with FCM levels in horses. Future research aims to determine the extent to which factors of influence on welfare, such as living conditions (e.g. single stalls versus group housing in pasture or paddocks), early life factors, and human interaction, act as mediators of cortisol levels in horses.
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Palme, R. (2019). Non-invasive measurement of glucocorticoids: Advances and problems. Physiol. Behav., 199, 229–243.
Abstract: Glucocorticoids (GCs; i.e. cortisol/corticosterone) are a central component of the stress response and thus their measurement is frequently used to evaluate the impact of stressful situations. Their metabolites from faeces of various animal species are more and more taken as a non-invasive aid to assess GC release and thus adrenocortical activity. The current literature review includes an extensive collection (1327 papers) and evaluation (see also Supplementary Tables) of the literature on faecal cortisol/corticosterone metabolite (FCM) analysis published to date. It aims at giving reference for researchers interested in implementing FCM analysis into their study or seeking to improve such methods by providing background knowledge on GC metabolism and excretion, conveying insights into methodological issues and stating caveats of FCM analysis and by highlighting prerequisites for and some examples of a successful application of such methods. Collecting faecal samples and analysing FCMs may appear simple and straightforward, but researchers have to select and apply methods correctly. They also need to be aware of the many pitfalls and potentially confounding factors and, last but not least, have to carefully interpret results. Applied properly, measurement of FCMs is a powerful non-invasive tool in a variety of research areas, such as (stress) biology, ethology, ecology, animal conservation and welfare, but also biomedicine.
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Hofmeester, T. R., Cromsigt, J. P. G. M., Odden, J., Andrén, H., Kindberg, J., & Linnell, J. D. C. (2019). Framing pictures: A conceptual framework to identify and correct for biases in detection probability of camera traps enabling multi-species comparison. Ecol Evol, .
Abstract: Abstract Obtaining reliable species observations is of great importance in animal ecology and wildlife conservation. An increasing number of studies use camera traps (CTs) to study wildlife communities, and an increasing effort is made to make better use and reuse of the large amounts of data that are produced. It is in these circumstances that it becomes paramount to correct for the species- and study-specific variation in imperfect detection within CTs. We reviewed the literature and used our own experience to compile a list of factors that affect CT detection of animals. We did this within a conceptual framework of six distinct scales separating out the influences of (a) animal characteristics, (b) CT specifications, (c) CT set-up protocols, and (d) environmental variables. We identified 40 factors that can potentially influence the detection of animals by CTs at these six scales. Many of these factors were related to only a few overarching parameters. Most of the animal characteristics scale with body mass and diet type, and most environmental characteristics differ with season or latitude such that remote sensing products like NDVI could be used as a proxy index to capture this variation. Factors that influence detection at the microsite and camera scales are probably the most important in determining CT detection of animals. The type of study and specific research question will determine which factors should be corrected. Corrections can be done by directly adjusting the CT metric of interest or by using covariates in a statistical framework. Our conceptual framework can be used to design better CT studies and help when analyzing CT data. Furthermore, it provides an overview of which factors should be reported in CT studies to make them repeatable, comparable, and their data reusable. This should greatly improve the possibilities for global scale analyses of (reused) CT data.
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Griebenow, K., & Klibanov, A. M. (1995). Lyophilization-induced reversible changes in the secondary structure of proteins. Proc Natl Acad Sci USA, 92(24), 10969–10976.
Abstract: Changes in the secondary structure of some dozen different proteins upon lyophilization of their aqueous solutions have been investigated by means of Fourier-transform infrared spectroscopy in the amide III band region. Dehydration markedly (but reversibly) alters the secondary structure of all the proteins studied, as revealed by both the quantitative analysis of the second derivative spectra and the Gaussian curve fitting of the original infrared spectra. Lyophilization substantially increases the beta-sheet content and lowers the alpha-helix content of all proteins. In all but one case, proteins become more ordered upon lyophilization.
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Palme, R., Touma, C., Arias, N., Dominchin, M.N., & Lepschy, M. (2013). Steroid extraction: Get the best out of faecal samples. Wien Tierärztl Monat – Vet Med Austria, 100, 238–246.
Abstract: Faecal steroid hormone metabolites are becoming increasingly popular as parameters for reproductive functions and stress. The extraction of the steroids from the faecal matrix represents the initial step before quantification can be performed. The steroid metabolites present in the faecal matrix are of varying polarity and composition, so selection of a proper extraction procedure is essential. There have been some studies to address this complex but often neglected point. Radiolabelled steroids (e.g. cortisol or progesterone) have frequently been added to faecal samples to estimate the efficiency of the extraction procedures used. However, native, unmetabolized steroids are normally not present in the faeces and therefore the results are artifi- cial and do not accurately reflect the actual recoveries of the substances of interest. In this respect, recovery experiments based on faecal samples from radiometabolism studies are more informative. In these samples, the metabolite content accurately reflects the mixture of metabolites present in the given species. As a result, it is possible to evaluate different extraction methods for use with faecal samples. We present studies on sheep, horses, pigs, hares and dogs that utilized samples containing naturally metabolized, 14C-labelled steroids. We recommend extracting faecal steroids by simply suspending the faeces in a high percentage of a primary alcohol (for glucocorticoid metabolites 80% aqueous methanol proved best suited for virtually all mammalian species tested so far). Not only does the procedure significantly increase the total amount of recovered radioactivity, it also increases the percentage of unconjugated metabolites, which are more likely to be recognized by the antibodies used in various immunoassays. The advantages of this extraction procedure are clear: it is very easy to use (no evaporation step is needed), it yields high recoveries and variation based on the extraction procedure is reduced to a minimum.
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Broekhuis, F., Madsen, E. K., & Klaassen, B. (2019). Predators and pastoralists: how anthropogenic pressures inside wildlife areas influence carnivore space use and movement behaviour. Anim Conserv, .
Abstract: Abstract Across the globe, wildlife populations and their behaviours are negatively impacted by people. Protected areas are believed to be an antidote to increasing human pressures but even they are not immune to the impact of anthropogenic activities. Areas that have been set aside for the protection of wildlife therefore warrant more attention when investigating the impact of anthropogenic pressures on wildlife. We use cheetahs Acinonyx jubatus as a case study to explore how a large carnivore responds to anthropogenic pressures inside wildlife areas. Using GPS-collar data we investigate cheetah space use, both when moving and stationary, and movement parameters (speed and turn angles) in relation to human disturbance, distance to human settlement, livestock abundance and livestock site use inside wildlife areas. Space use was negatively influenced by human disturbance, resulting in habitat loss and fragmentation and potentially reducing landscape permeability between neighbouring wildlife areas. Cheetahs were also less likely to stop in areas where livestock numbers were high, but more likely to stop in areas that were frequently used by livestock. The latter could reflect that cheetahs are attracted to livestock however, cheetahs in the study area rarely predated on livestock. It is therefore more likely that areas that are frequently used by livestock attract wild herbivores, which in turn could influence cheetah space use. We did not find any effects of people and livestock on cheetahs? speed and turn angles which might be related to the resolution of the data. We found that cheetahs are sensitive to human pressures and we believe that they could be an indicator species for other large carnivores facing similar challenges. We suggest that further research is needed to determine the levels of anthropogenic pressures needed to maintain ecological integrity, especially inside wildlife areas.
Keywords: cheetah; livestock; movement; human pressure; protected areas; space use
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Lee, P. (1991). Adaptation to environmental change:an evolutionary perspective. In H. O. Box (Ed.), Primate responses to environmental changes (pp. 39–56). London: Chapmann & Hall.
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Beck, B. B. (1980). Animal tool behaviour: The use and manufacture of tools by animals. New York: Garland.
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