Cortisol Enzyme Immunoassay Research Articles

Clippers are superior to scissors in the collection of hair for chemical analysis in companion dogs: a Dog Aging Project preliminary study.

To identify the safest, most efficient method for hair sample collection from companion dogs among clippers, scissors, and razors and to validate obtained samples with cortisol concentration analysis. 25 healthy, privately owned dogs. 2 hair samples were collected from each dog's ischiatic region with different implements (scissors, razors, or clippers). The collecting clinician completed a Hair Collection Questionnaire (HCQ) for each sample that compared subjective sample quality, time of collection, restraint needed, and patient experience. Each sample was evaluated by cortisol enzyme immunoassay. Clippers had higher overall HCQ scores than scissors, and scissors had higher HCQ scores than razors. Collection was faster for clippers than scissors, and scissors were faster than razors. There were no differences in sample quality between scissors and clippers, and sample quality was lower with razors. There was no difference in restraint needed or patient experience. Collection of long hair had higher HCQ scores than collection of medium and short hair. Collection of hair from dogs with an undercoat had higher HCQ scores than collection of hair from dogs without an undercoat. Dog size had no effect on HCQ score. Hair cortisol concentration did not vary between scissors or clippers (P = .111). Hair color and age did not affect hair cortisol concentration (P = .966 and P = .676, respectively). Clippers are recommended for hair sample collection from companion dogs. Scissors are an adequate alternative.

Analytical and biological validation of a noninvasive measurement of glucocorticoid metabolites in feces of Geoffroy's spider monkeys (Ateles geoffroyi).

We report on an analytical and biological validation of a commercial cortisol enzyme immunoassay (EIA)to measure glucocorticoids (GC)in feces of Geoffroy's spider monkeys (Ateles geoffroyi). Validation of endocrinological methods for each sample matrix and study species is crucial to establish that the methods produce reliable results. For the analytical validation of the EIA, we assessed parallelism, accuracy, and precision. We carried out a biological validation based on three well-studied GC patterns with the following predictions: (1) increased fecal GC metabolite (fGCM)concentrations after veterinary intervention; (2) increased fGCM concentrations during early morning hours; and (3) higher fGCM concentrations during gestation than in other female reproductive states. For the first prediction, we sampled feces of two zoo-housed females 2 days before, the day of, and 2 days after a veterinary intervention. For the second prediction, we analyzed 284 fecal samples collected from 12 wild males using a linear mixed model (LMM).For the third prediction, we analyzed 269 fecal samples of eight wild females using an LMM. Analytical validation revealed that the EIA showed parallelism, was accurate, and precise within each assay. However, there was elevated variation in between-assay precision. The biological validation supported all predictions: (1) the two zoo-housed females showed a substantial increase in fGCM concentrations 2.5and 11 h after veterinary intervention; (2) there was a negative effect of sample collection time on fGCM concentrations (i.e., higher concentrations during early morning); (3) gestating females had significantly higher fGCM concentrations than lactating females. Thus, we analytically validated the commercial EIA and, despite between-assay variation, we were able to find three biologically relevant GC signals in captive and wild settings, and in males and females. We are therefore confident that the method can be used to noninvasively address behavioral endocrinology questions in Geoffroy's spider monkeys.

Kibble diet is associated with higher faecal glucocorticoid metabolite concentrations in zoo-managed red wolves (Canis rufus).

The red wolf (Canis rufus) is a critically endangered canid that exists solely because of the establishment of the ex situ population in the late 1980s. Yet, the population under human care suffers from gastrointestinal (GI) disease in captivity. While the cause of GI disease is unknown, it is speculated that environmental factors can influence GI health of zoo-managed red wolves. The goal of the present study was to investigate the relationship between faecal glucocorticoid metabolite (FGM) concentrations, a biomarker for stress, and environmental factors for zoo-managed red wolves. Faecal samples were collected from 14 adult wolves three times a week for 5 to 12months. Using a single-antibody cortisol enzyme immunoassay, FGM concentrations were quantified. Environmental factors were collected for each participating wolf on dietary type, sex, type of public access to enclosure, density (enclosure size [ft2]/number of wolves living in enclosure) and a monthly average status of GI health. Red wolves that ate a commercial kibble diet had both higher FGM concentrations over time and higher baseline FGM concentrations compared to individuals that received commercial kibble mixed with commercial meat. Density, public access or GI health were not related to FGM concentration; however, males had higher baseline FGM concentrations compared to female red wolves. Our findings suggest that management conditions, particularly diet, can strongly influence FGM concentration in the zoo-managed red wolf population. Findings from this study highlight the importance of management choices on individual welfare. Maintaining a healthy captive population of red wolves is imperative for the persistence of the species, including successful future reintroductions.

Relationship between wool cortisol, wool quality indices of Australian Merino rams and climatic variables in Tasmania

IntroductionAustralia is the world’s largest producer and exporter of fine wool. External factors (i.e., non-genetic) that affect wool phenotype need to be identified and managed to minimize any negative effects on wool quality. Climatic stress is one external event that can affect wool phenotype, other stressors include shearing, nutrition, disease, pregnancy, and lambing. This study focuses on the relationship between Saxon Merino ram wool phenotype and wool cortisol. The hypotheses had two parts (i) wool cortisol levels would vary seasonally and variations would correlate with wool phenotype (ii) that these seasonal variations would also correlate with thermal indices.MethodsCortisol levels in Saxon Merino ram wool samples were determined using a cortisol enzyme-immunoassay (EIA) using polyclonal cortisol antibody. The wool cortisol results were correlated for significance with thermal indices including Temperature Humidity Index [THI] and Chill Index [CI] obtained from the Australian Bureau of Meteorology.ResultsPart 1 of hypothesis one was supported with wool cortisol showing significant seasonal variation. Part 2 of hypothesis 1 was not supported, with no statistically significant causality present between total wool cortisol levels and the price index of Saxon Merino ram wool. However, there was significant causality between wool cortisol and coarse edge measurement (CEM) of wool. Hypothesis 2, the proposal that thermal indices would impact on wool cortisol was also not supported.DiscussionIt can be concluded that although conclusive causality between total wool cortisol and wool phenotype quality could not be ascertained, there were indirect effects of cortisol levels on wool phenotypes with significant effects on the CEM. The CEM of Merino Saxon ram wool can be manipulated by assessing and managing cortisol levels during the growth cycles of Merino wool, through appropriate nutrition and husbandry procedures stress could be managed.

Measurement of Salivary Cortisol in Two New World Primate Species.

Glucocorticoids (GCs) are mammalian steroid hormones involved in a variety of physiological processes, including metabolism, the immune response, and cardiovascular functions. Due to their link to the physiological stress response, GC measurement is a valuable tool for conservation and welfare assessment in animal populations. GC levels can be measured from different matrices, such as urine and feces. Moreover, especially in captive settings, measuring GCs from saliva samples proved particularly useful as those samples can be collected non-invasively and easily from trained animals. Salivary GC levels can be measured using a variety of analytical methods, such as enzyme immunoassays. However, it is crucial to validate the analytical method for each specific application and species when using a new matrix. Using high-pressure liquid chromatography and a cortisol enzyme immunoassay, we show that the main glucocorticoids secreted in the saliva of squirrel monkeys and brown capuchin monkeys are cortisol and cortisone. Our biological validation found the expected salivary cortisol level to decline throughout the day. Our findings support the reliability of salivary cortisol measurements and their potential to be used as a valid tool in research and welfare assessment for these non-human primates.

Salivary cortisol response developed in Patients undergoing Dental treatment

Aims and objective: The stress control is essential in patients to elude the loss of motivation for Dental procedures .The present study was conducted to estimate salivary cortisol response to stress developed in patients undergoing Dental treatment Methodology : 36 patients had their salivary cortisol levels measured; 30 of them were in the study group and had had root canal, apicoectomy, tooth preparation, and impression-making procedures for the fixed prosthesis. Six of the patients were healthy individuals who didn’t need any dental treatment.The groups demarcated were as follows. Group 1: Control group with no treatment and just the saliva sample was taken. Group 2 ; The patients undergone access opening, Biomechanical preparation and obturation Group 3 ; The patients undergone apicoectomy procedure Group 4 ; The patients undergone tooth preparation and impression making for fixed prosthesis . The salivary cortisol levels were evaluated using Salivary Cortisol Enzyme Immunoassay Kit. Results : The salivary cortisol level was highest in Group 3,followed by Group 2 ,Group 4 and least in Group 1. Conclusion The stress level was lowest in healthy patient and highest when patient underwent apicoectomy procedure in appointment .

The Effect of Disease and Injury on Faecal Cortisol Metabolites, as an Indicator of Stress in Wild Hospitalised Koalas, Endangered Australian Marsupials.

Loss of habitat, urbanisation, climate change and its consequences are anthropogenic pressures that may cause stress in koalas. Non-invasive monitoring of faecal cortisol metabolites (FCMs) can be utilised to evaluate the impact of stressors. The aim was to determine if the tetrahydrocorticosterone (50c) and cortisol enzyme immunoassays (EIAs) could be effective in measuring FCM values in wild, stressed koalas. This research included 146 koalas from the Australia Zoo Wildlife Hospital (AZWH) and 88 from a study conducted by Endeavour Veterinary Ecology (EVE), Queensland, Australia. Faecal samples of diseased, injured and control koalas were analysed. The effect of hospitalisation on FCM values was also investigated. Diseased and injured koalas had significantly higher FCM values than clinically healthy control animals as measured by the 50c EIA. FCM values with the cortisol EIA differed significantly between control and diseased koalas, but not between control and injured ones. Moreover, only the 50c EIA detected higher absolute values in males compared to females, and also found that females showed a more elevated response to stress manifested by injury and disease compared to males. The 50c EIA detected stress during hospitalisation better than the cortisol EIA. The cortisol EIA was also found unreliable in detecting stress in rehabilitated koalas treated with synthetic glucocorticoids as it cross-reacts with these steroids providing artificially high values.

Evaluating Dermal Bone as a Novel Source of Endocrine Information in Ninespine and Threespine Stickleback Fish.

Monitoring the physiology of small aquatic and marine teleost fish presents challenges. Blood samples, often the first choice for endocrinologists, can be difficult or even impossible to obtain and alternative matrices currently used for hormone analyses do not occur in fishes (e.g., hair, feathers etc.) or are not easily collected from small aquatic organisms (e.g., urine and feces). Some teleosts, however, have enlarged bony dermal elements that possibly accumulate and store steroid hormones in physiological relevant concentrations. Both threespine stickleback (Gasterosteus aculeatus) and ninespine stickleback (Pungitius pungitius) have a series of external, lateral bony plates, dorsal spines, and a pair of pelvic spines attached to the pelvic girdle. We investigated if cortisol, the primary circulating glucocorticoid in teleosts, could be extracted from stickleback dermal bone and quantified using a commercially available enzyme immunoassay (EIA). We successfully validated a cortisol EIA for dermal bone extracts, determined that cortisol was detectable in both species, and found that dermal bone cortisol levels significantly correlated with cortisol levels in whole body homogenate. Ninespine stickleback had significantly higher dermal bone cortisol concentrations than threespine stickleback and female threespine stickleback tended to have over twice the mean dermal bone cortisol concentration than males. Because both stickleback species are widely used for ecotoxicological studies, using dermal bone as a source of endocrine information, while leaving the body for contaminant, genomic, histological, and stable isotope analyses, could be a powerful and parsimonious tool. Further investigation and physiological validations are necessary to fully understand the utility of this new sample matrix.

Glucocorticoid concentration and parasitic load in a wild striped hyaena (Hyaena hyaena) population in Southern India

Glucocorticoid levels (GCs) are an important parameter that infers the pathological condition of an organism. GCs can vary with life history stages like gender, age, and reproductive stage and signify the degree of allostatic load experienced. Deleterious increases in GCs can severely affect the wildlife population by suppressing the reproductive viability of individuals and may cause immunosuppressive, making the population more susceptible to parasitic load. We have studied the stress physiology and parasitic load across the life history stages in the striped hyaena population confined to the Moyar Valley of Mudumalai and Sathyamangalam Tiger Reserves. The fecal glucocorticoid metabolite concentrations were measured using cortisol enzyme immunoassay from 30 scats. Collected scats were categorized into adults, sub-adults, and juvenile age groups based on the size and colour of droppings. Sex of the scat samples was identified post-DNA extraction from their epithelial lining cells. Parasitic load from scats was quantified as the number of different parasites recorded through sucrose floatation and sedimentation techniques. We found no significant difference in GCs concentration and parasitic load across the life history stages, but juveniles were recorded to have the least parasitic load. The highest GCs concentration and parasitic load were recorded from adults and female individuals, while both parameters were found least among juvenile individuals. Though we did not observe any significant differences across life-history stages, this study provides the first baseline information on stress physiology and parasitic load in striped hyaenas, which could help further studies assess the impact of external perturbations on stress physiology.

Body weight loss after birth increases stress in the newborns

BackgroundBody weight loss is physiological when is lower than 8–10% in the first days of life. Low birth body weight is correlated to up regulation of hypothalamus-pituitary-adrenal axis function. The aim was to test the hypothesis that weight loss from 24 to 48 h of life is associated with high levels of salivary cortisol in healthy term newborns. Methodsa prospective clinical study was performed enrolling healthy term newborns who started the rooming- in the delivery room to improve skin to skin and breastfeeding. Salivary cortisol levels were measured at 24 and 48 h of life, using Salimetrics Salivary Cortisol Enzyme Immunoassay Kit. Weight loss/gain was recorded every 24 h starting from birth. ResultsA significant positive correlation was found between salivary cortisol levels and body weight loss at 48 h of life (n = 57, r = 0,393; p = 0,006). At 48 h of life, salivary cortisol levels were significantly higher in newborns that lost body weight 1,7 times more than body weight loss at 24 h of life compared to newborns who maintained a more stable body weight between 24 and 48 h of life ConclusionsThe swift body weight loss from 24 to 48 h of life increases salivary cortisol levels in term newborns. Data suggest the need to protect exclusive breast fed newborns with human milk donor in term newborns, in case of temporary breast milk lack, to reduce cortisol exposure early in life.

Preliminary Investigation into Developing the Use of Swabs for Skin Cortisol Analysis for the Ocean Sunfish (Mola mola).

Simple SummaryZoos and aquaria play an important role in preventing the mass extinction of wildlife through public awareness of conservation issues and providing a safe haven for wildlife populations. Because aquatic populations face many challenges due to pollution and global warming, it is important to develop an understanding of how species can cope with their environment whether it be in the wild or under human care. Here, we were interested in developing non-invasive methods to study fish stress physiology. We use the unique ocean sunfish (Mola mola) to develop the use of skin swabs to measure the stress hormone, cortisol. We used known times of stress including when a mola was injured or ill and during acclimation to the Monterey Bay Aquarium. We found that cortisol increased initially within the first month of being admitted to the aquarium, but returned to normal values afterward. Molas also had elevated cortisol when being treated for an injury or illness. This is the first step in the development of the use of skin swabs to collect samples for stress analysis in the mola. Additional biochemical analysis is needed to confirm these results and allow this method to be applied to other species of fish.The ocean sunfish (mola; Mola mola) is the heaviest bony fish in the world. This slow-moving fish often is injured by fishing boats that use drift gillnets attributing to its listing as Vulnerable by the IUCN. The Monterey Bay Aquarium (Monterey, CA, USA) has a program that brings in smaller molas from the ocean and acclimates them for an exhibit. When they grow too large for the million-gallon Open Seas exhibit, they are returned to Monterey Bay through a “reverse” acclimatization. Our overall goal was to use skin swabs to evaluate mola stress physiology to better understand the effects of this program. Our objectives were to validate this non-invasive method by taking opportunistic swabs throughout acclimatization and during stressful events. We swabbed each individual (n = 12) in three different body locations. Swabs were analyzed using a cortisol enzyme immunoassay. We averaged the three swabs and examined the absolute change of cortisol from the first taken upon handling to during treatments and the different acclimation stages. We considered elevated cortisol concentrations to be ≥1.5-fold higher than the first sample. Overall, mean (±SEM) cortisol varied among individuals (564.2 ± 191.5 pg/mL swab (range, 18.3–7012.0 pg/mL swab). The majority (four of six) of molas swabbed within the first week or month had elevated skin cortisol compared to their first sample. All seven molas that were being treated for an injury or illness had elevated skin cortisol (range, 1.7- to 127.6-fold higher) compared to their post-acclimation sample. This is the first step in validating the use of non-invasive skin swabs for glucocorticoid analysis in the mola. Further biochemical analysis is needed to determine the specific steroids that are being measured.

Intra- and Inter-sample Variation in Wool Cortisol Concentrations of Australian Merino Lambs Between Twice or Single Shorn Ewes

Stress in Merino sheep can cause a reduction in the quantity and quality of fine wool production. Furthermore, it has been found that environmental stress during pregnancy can negatively affect the wool follicles of the developing fetus. This study was part of a larger field investigation on the effects maternal shearing frequency on sheep reproductive and productivity outcomes. For this study, we investigated the intra- and inter- sample variation in wool cortisol levels of weaner lambs. We conducted two experiments, the first was to determine the intra- and inter- sample variation in wool samples taken from the topknot of weaned lambs, and the other aim was to determine any difference between maternal shearing treatment (single or twice shearing) on absolute wool cortisol levels of weaned lambs. In the first experiment, topknot wool was collected from 10 lambs, and each sample was further divided into four subsamples, leading to a total of 40 wool subsamples. For the second experiment, we collected the topknot from the 23 lambs produced by the shearing frequency treatment ewes (once or twice shorn). The samples were then extracted and analyzed using a commercially available cortisol enzyme-immunoassay in order to determine the concentration of cortisol in each of the samples. Statistical analysis for the first experiment showed that there was no significant difference between the subsamples of each topknot wool sample taken from each lamb (p = 0.39), but there was a statistical difference between samples (p < 0.001), which was to be expected. In the second experiment, there was a significant difference between the lambs born to the one shearing and two shearing treatments (p = 0.033), with the lambs of the twice sheared ewes having higher average wool cortisol levels [2.304 ± 0.497 ng/g (SE); n = 14] than the ones born to once shorn ewes [1.188 ± 0.114 ng/g (SE), n = 8]. This study confirms that the topknot wool sampling can be a reliable method adapted by researchers for wool hormonal studies in lambs. Second, ewes shorn mid-pregnancy gave birth to lambs with higher cortisol concentrations than ewes that remained unshorn during pregnancy. This result warrants further investigation in a controlled study to determine if maternal access to nutrition (feed and water) may impact on the HPA-axis of lambs.

Fecal Glucocorticoid Metabolites as a Noninvasive Indicator of Stress in the Tsushima Leopard Cats (Prionailurus bengalensis euptilurus): Application to Health Care.

Simple SummaryThe Tsushima leopard cat (Prionailurus bengalensis euptilurus) and the Iriomote cat (P. b. iriomotensis) are the only two wild felids living in Japan. Both species are endangered. The Tsushima leopard cat is undergoing captive breeding (ex situ conservation) in zoos with the aim of returning it to the wild, in addition to conservation activities in the habitat (in situ conservation). In order to promote captive breeding, it is important to elucidate the reproductive physiology and to evaluate stress conditions. We investigated the possibility of measuring glucocorticoid metabolites (GCMs), which are stress hormones, using feces in Tsushima leopard cats. We examined changes in fecal GCM in individuals who underwent a health examination under anesthesia as an unavoidable stressful condition during normal health care. Fecal GCM concentrations increased 1–2 days after the health examination. Fecal GCM concentrations were higher in diseased individuals than in apparently healthy individuals. Individuals diagnosed with disk herniation also showed a decrease in fecal GCM concentrations after treatment with therapeutics. These results indicate that the measurement of fecal GCMs is useful for improving the husbandry and health management of the Tsushima leopard cat.This study investigates whether the measurement of glucocorticoid metabolites (GCMs) in feces is a useful method for the noninvasive evaluation of stress in the endangered Tsushima leopard cats (Prionailurus bengalensis euptilurus). Feces were collected from six seemingly healthy and five diseased (renal dysfunction, adrenal tumor, hernia, feline immunodeficiency virus (FIV), feline leukemia virus (FeLV)) Tsushima leopard cats in captivity. Fecal GCMs were measured by enzyme immunoassay (EIA) for cortisol. Individuals that experienced a physical examination under anesthesia showed increased fecal GCMs 1–2 days after the event. An individual diagnosed with disk herniation showed decreased fecal GCMs after medical administration. The mean fecal GCM concentrations for six healthy animals and five diseased animals were 0.66 ± 0.08 and 2.65 ± 0.76 μg/g, respectively, which was significantly different. Cortisol and corticosterone were not clearly detected in the feces examined by the use of the HPLC-EIA analysis. GCMs may be excreted in the feces; however, the exact identification of these substances is not achieved. The results suggest that the measurement of fecal GCMs is useful for the husbandry and health management of this species.

Over Time Decay of Cortisol Metabolites in Faecal Pellets of Koalas in Central Queensland.

Simple SummaryFaecal cortisol metabolites (FCMs) are a useful, non-invasive tool for the assessment of stress in koalas. However, FCM stability after defecation is a critical issue. Therefore, we exposed faecal pellets of koalas to three different environmental conditions and measured FCMs with three recently evaluated enzyme immunoassays (EIAs). Because water loss over time exerted the greatest influence on FCMs, we strongly recommend collecting freshly defecated pellets in koalas.Faecal material can be a valuable source of information for a range of animal health aspects and can be used to measure faecal cortisol metabolites (FCMs). FCM values can relate to physiological stress responses. However, freshly defecated pellets are not always available and environmental conditions, such as temperature and humidity, might affect faecal pellet consistency and FCM levels. Therefore, the impact of environmental conditions on FCMs needs to be evaluated. We collected 107 pellets from two female and two male koalas, exposed them to three types of treatments, and analysed FCMs in these samples with three enzyme immunoassays (EIAs). After analysis, the original FCM values were mathematically corrected for water loss. Results show that the FCMs were more stable when measured using tetrahydrocorticosterone (50c) and 5α-pregnane-3β,11β,21-triol-20-one (37e) EIAs, and were less stable when measured with the cortisol EIA. With 50c, the FCM values did not vary significantly over time either before or after the adjustment with water in the environment treatment group. For samples kept under constant low (25 °C) and high (35 °C) temperatures, the 50c FCM values did not vary significantly over time, after adjustments were made for water loss. Thus, this study highlights the importance of considering the suitability of faecal field samples for FCM analysis. Because water loss was the main driver of FCM changes, we strongly recommend collecting koala pellets that are freshly defecated, despite the effort and time it might take to collect such pellets.

Reference intervals and values for fecal cortisol, aldosterone, and the ratio of cortisol to dehydroepiandrosterone metabolites in four species of cetaceans.

The goal of the current study was to create reference intervals and values for several common and one potential novel physiological indicators of animal welfare for four species of cetaceans. The subjects included 189 bottlenose dolphins (Tursiops truncatus), 27 Indo-Pacific bottlenose dolphins (Tursiops aduncus), eight Pacific white-sided dolphins (Lagenorhynchus obliquidens), and 13 beluga whales (Delphinapterus leucas) at Alliance of Marine Mammal Parks and Aquariums and/or Association of Zoos and Aquariums accredited facilities. During two sampling time periods between July and November of 2018 and between January and April of 2019, fecal samples were collected weekly for five weeks from all animals. Samples were processed and analyzed using enzyme immunoassay for fecal cortisol, aldosterone, and dehydroepiandrosterone (DHEA) metabolites. Linear mixed models were used to examine demographic and time factors impacting hormone metabolite concentrations. Age, sex, and time of year were all significant predictors for some of the models (p < 0.01). An iOS mobile application ZooPhysioTrak was created for easy access to species-specific reference intervals and values accounting for significant predictors. For facilities without access to this application, additional reference intervals and values were constructed without accounting for significant predictors. Information gained from this study and the use of the application can provide reference intervals and values to make informed management decisions for cetaceans in zoological facilities.

Hair Cortisol, Testosterone, Dehydroepiandrosterone Sulfate and Their Ratios in Stallions as a Retrospective Measure of Hypothalamic-Pituitary-Adrenal and Hypothalamic-Pituitary-Gonadal Axes Activity: Exploring the Influence of Seasonality.

Simple SummaryThe monitoring of a horse’s endocrine status can provide valuable information of its welfare and sexual and health conditions. The use of non-invasive matrixes for hormonal monitoring, as alternatives to blood samples, are being increasingly used. Among them, the measurement of hormones deposited in the hair shaft has been widely used as a retrospective hormonal biomarker. In the present study, we aimed to evaluate the seasonal variations in hair C, T and DHEA-S in horses for a whole year, as well as to assess the variations between seasons of C/DHEA-S and T/C ratios as a retrospective measure of the hypothalamic–pituitary–adrenal and hypothalamic–pituitary–gonadal axis activity. The results showed how cortisol, testosterone and dehydroepiandrosterone sulphate and their ratios were significantly affected by season. Therefore, as shown in this study, season should be considered when analysing sexual and stress hormones in stallion hair.The monitoring of stress physiology includes studying a wide range of endocrinological mechanisms, which can be assessed using multiple tissue samples. This study aimed to evaluate the seasonal variations of hair C, T and DHEA-S in horses for a whole year, as well as to assess the variations between seasons of C/DHEA-S and T/C ratios as a retrospective measure of the hypothalamic–pituitary–adrenal and hypothalamic–pituitary–gonadal axis activity. Ten pure-breed Menorca stallions were included in the study. The hair samples were collected approximately every two months following the shave-reshave method caudally to the sternum. After a methanol-based extraction, samples were analyzed by enzyme immunoassay for cortisol, testosterone, and dehydroepiandrosterone sulphate. Following our findings, we detected that cortisol, testosterone and dehydroepiandrosterone sulphate were significantly affected by seasonality, with the highest values of cortisol during summer and the lowest values of testosterone during spring. Dehydroepiandrosterone sulphate concentrations were increased in autumn compared to the other studied periods. Additionally, the studied hormone ratios showed variations between seasons. To conclude, season should, therefore, be considered when assessing sexual and stress hormones in stallion hair, since this variable can be a potential influencing factor and led to misinterpretations.

Validating the use of fecal glucocorticoid metabolite analysis to assess the adrenocortical activity of the zoo-housed Sichuan takin (Budorcas taxicolor tibetana).

With limited information known about the zoo-housed Sichuan takin (Budorcas taxicolor tibetana), there is a need to gain more knowledge about their basic physiology to be able to better assess their well-being. Our goal was to develop noninvasive methods to evaluate stress physiology in theSichuan takins housed under human care. The objectives were: (1) validate the use of fecal glucocorticoid metabolite (FGM) analysis to monitor adrenocortical activity and (2) determine the relationship between FGM concentrations and changes in various factors including environmental conditions, reproductive hormones, and social factors. Three (one male and two females) adult Sichuan takins were included in the study from Lincoln Park Zoo. A cortisol enzyme immunoassay was used to analyze FGMfrom samples that were collected 2-4 times per week. FGMwas biochemically validated in the laboratory and biologically validated using pregnancy and parturition. Results showed that 1 day after birth one female had a more than a six-fold increase in FGM. A positive relationship (p < .05) was observed between FGM and temperature for twoof the threetakins. Finally, FGM values tended to increase during times when aggression (p = .09) and reproductive (p = .08) behaviors were observed. In conclusion, environmental, reproductive, and behavioral factors could affect adrenocortical activity in zoo-housed Sichuan takins; therefore, these methods can be used to assist in the management and conservation of this threatened species both in zoos and potentially in the wild.

Correcting for enzyme immunoassay changes in long term monitoring studies

Enzyme immunoassays (EIAs) are a common tool for measuring steroid hormones in wildlife due to their low cost, commercial availability, and rapid results. Testing technologies improve continuously, sometimes requiring changes in protocols or crucial assay components. Antibody replacement between EIA kits can cause differences in EIA sensitivity, which can hinder monitoring hormone concentration over time. The antibody in a common cortisol EIA kit used for long-term monitoring of stress in wildlife was replaced in 2014, causing differences in cross reactivity and standard curve concentrations. Therefore, the objective of this study was to develop a method to standardize results following changes in EIA sensitivity. We validated this method using cortisol concentrations measured in the hair of brown bears (Ursus arctos).•We used a simple linear regression to model the relationship between cortisol concentrations using kit 1 and kit 2.•We found a linear relationship between the two kits (R2 = 0.85) and used the regression equation (kit2 = (0.98 × kit1) + 1.65) to predict cortisol concentrations in re-measured samples.•Mean predicted percent error was 16% and 72% of samples had a predicted percent error <20%, suggesting that this method is well-suited for correcting changes in EIA sensitivity.

Assessment of the changes in the stress related salivary cortisols levels induced during different Dental procedures

Aims and objective: Anxiety ,fear and pain assessment is essential in day to day Dental practice .The present study evaluated the stress related salivary cortisols levels induced during scaling,tooth preparation and surgical flap elevation in Periodontally compromised subjects &#x0D; Methodology: The salivary cortisol level were assessed in 80 patients ,out of which 40 patients were healthy patient who do not require any Dental treatment and 40 patients who underwent Scaling , tooth preparation and surgical flap elevation in Periodontally compromised subjects &#x0D; The groups divided were as follows .&#x0D; Group 1: control group with no treatment and just the saliva sample was taken.&#x0D; Group 2; The patients underwent scaling&#x0D; Group 3; The patients underwent tooth preparation for fixed prosthesis&#x0D; Group 4; The patients underwent surgical flap elevation in Periodontally compromised situation .The salivary cortisol levels were assessed using Salivary Cortisol Enzyme Immunoassay Kit.&#x0D; Results: The salivary cortisol level was highest in Group 4 followed by Group 2, Group 3 and least in Group 1.&#x0D; Conclusion: The stress level was lowest in healthy patient and highest when patient underwent surgical flap elevation in Periodontally compromised subjects .Dentist should try to minimize subject anxiety and stress to great extent .

Non-invasive monitoring of adrenocortical function in female domestic pigs using saliva and faeces as sample matrices.

Intensive pig management involves in a commercial setting the housing and implementation of certain procedures, such as castration and tail docking, which may be stressful for the animal. Good farming practices include the reduction of stress due to management processes, but assessing the level of stress perceived entirely through behavioural observations can be challenging. The monitoring stress-related physiological markers, like glucocorticoids (GC), can be an accurate alternative that would presumably be more objective. In order to avoid an additional stressor by taking blood, a non-invasive approach is advisable. We used an adrenocorticotropic hormone (ACTH) stimulation test and the effect of transport to examine the suitability of different enzyme immunoassays (EIAs) for monitoring adrenocortical function in domestic pigs using saliva and faeces as sample matrices. An assay measuring faecal glucocorticoid metabolites (fGCMs) with a 3ß,11ß-diol group has proven suited to determine adrenocortical activity, showing an overall increase of 180% in fGCM concentrations related to ACTH administration and of 70% related to transport, respectively. A cortisol EIA was used to detect salivary glucocorticoid (sGC) concentrations, revealing a 1100% increase in sGC concentrations after ACTH administration. The stability of fGCM concentrations post-defecation was determined to assess possible changes in measured fGCM concentrations in unpreserved faecal material over time, with fGCM concentrations being relatively stable (maximal 12% change) under natural conditions for approximately two days after defecation. This implicates that untreated faecal material from pigs can be analysed for up to two days after collection without appreciable level of depreciation in fGCM concentrations. Being able to assess the physiological stress response of domestic pigs non-invasively can help to improve the well-being of commercially reared pigs.