Non-invasive hormone assessment of Australian Merino Rams (Ovis aries): A pilot investigation of cortisol, testosterone and heat stress

Non-invasive hormone assessment is growing in interest as producers and livestock researchers seek new methods to assess animal welfare. Conventional matrices used for hormone assessment include blood serum, saliva, urine and faeces, typically involving invasive human-animal interaction, which is known to initiate an acute stress response and thus confound studies assessing cortisol. As such, these matrices are most appropriate as short-term, point measures as they reflect circulating concentrations at the level of the minute and hour. Alternatively, non-invasive hair and wool assessment offer long-term, historic reflections of hormone concentration at the scale of weeks and months - and are not limited by sampling stress - thus making wool an appropriate tissue for hormone analysis. This pilot study quantified cortisol and testosterone concentrations of ram fleece and determine if there is a significant difference between segments of the sample staple, and whether there is a correlation between hormones. Cortisol is a glucocorticoid produced within the adrenal glands and secreted in anticipation of or in response to a stressor. Testosterone is an androgen mainly synthesised within the testes of males and responsible for several critical functions including regulation of muscle growth, libido and spermatogenesis. In our study, 70 topknot wool samples were collected from rams on a commercial stud property in Dirranbandi, Queensland, Australia. Of these animals, 12 samples were selected at random to undergo cortisol and testosterone quantification. In the laboratory, a single, intact staple was isolated from the total sample, divided into 10 mm segments and prepared for their respective (cortisol or testosterone) immunoassays. No significant difference (p>0.05) was found between wool segments for either cortisol or testosterone, however, statistical differences (p<0.05) were found between individuals for both hormones. A strong correlation (R2=0.9173) was found between wool cortisol and testosterone concentrations, which was a first-time discovery in rams. Furthermore, climate loggers (n=6) were installed in proximity to the rams to collect daily maximum temperature ({degrees}C) and relative humidity (RH%) data to calculate the temperature-humidity index (THI) as an indicator of potential heat stress. Under this method, rams were deemed comfortable at a THI<72; under mild stress at a THI of between 72 and 78; moderate stress between 79 and 80; and severe stress at a THI of >81. Mean weekly THI peaked in late summer (February), remained high in early autumn (March), decreased throughout the remainder of autumn and the duration of winter before increasing slightly as temperatures rose in early spring (September). Over the trial, 90.36% or 4,706 h were marked by comfortable conditions, 8.95% or 466 h by mild stress, 0.60% or 31 h by moderate, and 0.10% or 5 h by severe stress. It was determined that THI is most valuable when combined with other methods of measuring heat stress, including non-invasive wool hormone assessment. Whilst most of the findings in this study were previously confirmed by other studies, the strong correlation between wool cortisol and testosterone concentrations appears to be a first. In summary, this study reveals the major future possibilities for non-invasive wool hormone assessment and the possible applications of combining this with temperature-humidity index scores to provide further insight into heat stress within the context of production animal agriculture.