Subject(s)
Hydrocortisone , Phascolarctidae , Animals , Hydrocortisone/metabolism , Phascolarctidae/metabolism , FecesABSTRACT
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.
ABSTRACT
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-3ß,11ß,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.
ABSTRACT
The koala (Phascolarctos cinereus) is an arboreal folivorous marsupial endemic to Australia. Anthropogenic activities and climate change are threats to this species' survival and are potential stressors. A suitable non-invasive method is needed to objectively detect stress in koalas. Under conditions of stress, the concentration of the hormone cortisol in plasma or in saliva is elevated, and this would provide a convenient measure; however, collecting blood or saliva from wild animals is both practically difficult and stressful, and so likely to confound any measurement. In contrast, measurement of cortisol metabolites in faeces provides a practical and non-invasive method to objectively measure stress in koalas. Unfortunately, the identity of the main faecal cortisol metabolites of koalas is unknown. In this study, we have used both untargeted liquid chromatography-mass spectrometry (LC-MS) and enzyme immunoassays (EIAs) to identify several faecal cortisol metabolites in two koalas, one female (18 months old, 4.1 kg) and one male (4 years old, 6.95 kg) upon administration of hydrocortisone (cortisol) sodium succinate. The LC-MS analysis identified tetrahydrocortisol along with several other isomers as cortisol metabolites. After a survey of five enzyme immunoassays, we found that two metabolites, tetrahydrocortisol and 3ß-allotetrahydrocortisol, could be detected by EIAs that used antibodies that were raised against their structurally similar corticosterone counterparts, tetrahydrocorticosterone and 3ß-allotetrahydrocorticosterone, respectively. While the 3ß-allotetrahydrocortisol metabolite was detected in the faeces of only one of the two animals studied, tetrahydrocortisol was detected in both. These results ultimately indicate that tetrahydrocortisol is likely the main faecal cortisol metabolite in koalas, and we demonstrate that it can be measured by an EIA (50c) that was originally developed to measure tetrahydrocorticosterone.
ABSTRACT
The Koala (Phascolarctos cinereus) is an endemic marsupial inhabiting four states of Australia. Urbanisation, declining habitat, drought and fires are threatening the survival of this flagship species. These threats may cause acute and chronic stress in koalas, which might also be associated with occurrence of infectious diseases in koala populations. Stress may induce an increase in cortisol reflected in increased faecal cortisol metabolite (FCM) values. To be able to use faecal cortisol metabolites to measure stress levels in this species, our aim was to determine baseline values for males and females during breeding and non-breeding season. A total of 351 defecations were collected fortnightly, twice a day, for 12 months from koalas at a wildlife facility in South East Queensland. Samples were analysed with three different enzyme immunoassays (EIAs): a cortisol, 5α-pregnane-3ß,11ß,21-triol-20-one (37e) and tetrahydrocorticosterone (50c) EIA. The latter, which also reacts with tetrahydrocortisol, the main metabolite in koala faeces, was found to have the highest biological sensitivity and, therefore, is the most suitable EIA to measure stress levels in koalas. Utilising this EIA, we found significant differences (p < 0.05) in FCM values between males and females, breeding and non-breeding season, and between morning and evening samples. Values of faecal cortisol metabolites established in stress-free koalas in this study can serve as a reference for future studies in koalas.
