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

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-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.

SUBMITTER: Santamaria F

PROVIDER: S-EPMC8698095 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Welfare studies of blue foxes would benefit from a measurement of faecal cortisol metabolites (FCMs) as a non-invasive, physiological stress parameter reflecting hypothalamus-pituitary-adrenal (HPA) axis activity. Before implementation, a species-specific validation of such a method is required. Therefore, we conducted a physiological validation of an enzyme immunoassay (EIA) to measure FCMs in blue foxes. Twenty individuals (nine males and eleven females) were injected with synthetic adrenocorticotrophic hormone (ACTH) and faecal samples were collected every third h for two days. The FCM baseline levels were assessed based on the first sampling day (control period, 144 samples), followed by the ACTH injection and the second day of sampling (treatment period, 122 samples). FCMs were analysed with a 5α-pregnane-3ß,11ß,21-triol-20-one EIA. We compared the estimated mean FCM concentrations of the treatment samples to the baseline average. All samples for the two periods were collected at the same time of the day, which enabled to test the data also with an hourly pairwise comparison. With the two statistical approaches, we tested whether a possible diurnal fluctuation in the FCM concentrations affected the interpretation of the results. Compared to the baseline levels, both approaches showed 2.4-3.2 times higher concentrations on time points sampled 8-14 h after the ACTH injection (p < 0.05). The estimated FCM concentrations also fluctuated slightly within the control period (p < 0.01). Inter-individual variations in FCM levels were marked, which highlights the importance of having a sufficient number of animals in experiments utilising FCMs. The sampling intervals of 3 h enabled forming of informative FCM curves. Taken together, this study proves that FCM analysis with a 5α-pregnane-3ß,11ß,21-triol-20-one EIA is a valid measurement of adrenocortical activity in the farmed blue foxes. Therefore, it can be utilised as a non-invasive stress indicator in future animal welfare studies of the species.

Project description:Highlights • Dried urine is an alternative to liquid urine for measuring cortisol and its metabolites.• 4-spot urine sampling is representative of a 24-hour urine collection.• 4-spot urinary cortisol measures reflect the diurnal pattern of change in cortisol.• Comprehensive profiling of cortisol is viable using both saliva plus dried urine samples. Aims/objectives The primary aim of this study was to determine the utility of dried urine sampling in obtaining measures of cortisol and cortisol metabolites. Additional aims were to evaluate if a 4-spot dried urine collection is representative of a 24-hour urine collection and if expected diurnal cortisol patterns can be observed in samples from both urine and saliva. Methods Data from individuals with cortisol measures available from both a 4-spot dried urine collection and a 24-hour urine collection (n = 28) were evaluated. Of these 28, 20 also had concurrent liquid and dried 24-hour urine measures. Consistency between these methods was evaluated using paired t-tests and intraclass correlation coefficients (ICCs). In addition, data from individuals with concurrent measures of both urinary and salivary cortisol (n = 68) were assessed for consistency in the diurnal pattern of change in cortisol. Results Near ideal consistency was observed between liquid and dried urine for measures of total urine free cortisol, total urine cortisone, and total cortisol metabolites (n = 20; ICCs = 0.99, 0.97 and 0.96, respectively). Good to excellent consistency was observed between the 4-spot method and the 24-hour collection (n = 28; ICCs = 0.89, 0.95 and 0.92, respectively). In mixed model analysis, no difference was seen in the diurnal pattern of cortisol between salivary and urinary free cortisol (n = 68; P = 0.83). Conclusion Dried urine is a viable alternative to liquid urine for the measurement of cortisol and cortisol metabolites. Additionally, if the 4 measures are added together, 4-spot urine collections can be representative of diurnal cortisol patterns commonly assessed using saliva and 24-hour urine collections.

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

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