Captive breeding programs are increasingly being used as a management option for threatened mammals. The greater bilby Macrotis lagotis, for example, is a vulnerable species which is maintained in captivity at several facilities in Australia. Non-invasive evaluation of stress hormones (cortisol in mammals) via excretory metabolites can be used to monitor physiological stress responses of captive individuals. In this study, we validated an enzyme-immunoassay (EIA) to measure cortisol metabolites in fresh faecal samples of adult male and female bilbies (n = 7) held in captivity at the Dreamworld Theme Park, Queensland, Australia. The faecal cortisol EIA was validated via parallelism and the recovery of exogenous cortisol added to pooled faecal extracts (> 99% recovery). Female bilbies had higher average faecal cortisol metabolite concentrations and higher day-today variation than male bilbies; however, there was no relationship with bilby age. Cortisol metabolites for most individuals varied widely through time, with numerous peaks and troughs in response to long-term stressors (illnesses, injury and reproductive issues) and short-term stressors, such as use in shows at Dreamworld or public displays in local schools, manual restraint and short-term veterinary procedures (e.g. general anaesthesia). Overall, the higher mean cortisol metabolite concentrations of individuals suffering long-term stress was related to a greater response to short-term stressors. This suggests an interaction between responses to short-term and long-term stressors which is perhaps due to habituation and/or facilitation of long-term stressors. Non-invasive faecal monitoring of stress hormones could provide further information on the implications of captive breeding programs and the release of animals reared in captivity.

Natural weathering conditions can influence faecal cortisol metabolite (FCM) measurements in wildlife if fresh faeces cannot be collected immediately following defaecation. In this study, we evaluated this issue in a threatened Australian marsupial, the greater bilby (Macrotis lagotis). Fresh (<12 h since defaecation) faecal samples (n = 19 pellets per bilby) were collected one morning from seven adult bilbies kept in captivity. One control faecal sample (Day 1) from each bilby was immediately frozen. The remaining faecal pellets were randomly positioned outdoors. Subsequently, we froze one faecal pellet every 24 h for 19 days. FCM levels in bilby faeces were quantified using an enzyme-immunoassay. Mean FCM levels showed variation (daily mean coefficients of variation [CV %]) of 56.83–171.65% over 19 days. Overall, FCM levels were affected by exposure time; however, multiple comparisons showed that no significant change in FCM occurred after environmental exposure (no significant difference in mean FCM between control (Day 1) with any of the exposure days (Days 2–19). Individuals and sex also affected FCM levels. We found no correlation between mean daily CVs with daily minimum–maximum temperatures or rainfall. Our results indicate that FCM in bilby faeces is fairly stable to long-term environmental exposure (19 days). In future, freshly excreted bilby faeces (where the sample maintains a distinct odour for 9–13 days) should be used to study FCM levels in wild bilbies.

Rapid and reliable physiological evaluation of stress is necessary for understanding the potential impacts of environmental changes on managed populations of threatened mammals. In situ populations of Australia's iconic marsupial, the greater bilby (Macrotis lagotis), are nearing extinction due to the impacts of competition and predation by feral animals and unpredictable climatic events (summer heat waves). In this study, we focussed our aim to identify a non-invasive method to measure adrenal activity in the species and also to identify potential factors that should be considered when comparing physiological stress in semi-free ranging populations of the species. We validated an enzyme immunoassay (EIA) for detecting fecal cortisol metabolites (FCM) from fresh fecal pellets taken from bilbies within four captive sites and two semi-free ranging populations around Queensland and New South Wales, Australia. Our FCM EIA successfully detected the 'raise and fall' pattern of FCM levels within 3 days of exogenous adrenocorticotropic hormone (ACTH) challenge. Mean FCM levels differed significantly between the captive sites and between sexes. All male bilbies grouped outdoor in captivity expressed the highest mean FCM level in comparison to all captive males that were housed individually or as groups indoors. Also, semi-free ranging bilbies expressed higher mean FCM levels than the captive bilbies. Overall, our study successfully validated a non-invasive tool for monitoring physiological stress in the greater bilby. In the future, it will be worthwhile to consider factors such as housing conditions, sex and location when comparing the adrenal sensitivity to environmental changes, to help evaluate the success of management interventions (such as predator free enclosures) and support the survival of the species.

Small mammal trapping is a commonly taught field technique in ecology and population biology. However, there are still open questions regarding the extent to which trapping affects the welfare of small animals. This study investigated faecal cortisol as a non-invasive measurement of the physiological stress response of bank voles (Myodes glareolus) to live-trapping. Faecal cortisol concentrations were similar between samples collected from trapped and non-trapped voles, but were negatively correlated with capture duration. This result may suggest that any physiological stress response was short-lasting and the animals habituated to the trap. As such, effects on faecal cortisol were not apparent as mean faecal cortisol concentration was determined from multiple collected faeces. Future work should focus on characterising the potential stress response to trapping with greater temporal resolution.