Leopards are versatile predators. These elusive cats can successfully occupy any habitat that supports sufficient numbers of prey species and which provides adequate cover for their ambush-style of hunting.

Leopards also adapt well to settled environments near human activity. But this often brings them into conflict with humans. In South Africa it's been clear since the late 1980s that although protected areas play an important role in leopard conservation, most of the country's suitable leopard habitat lies outside the boundaries of protected areas, often on private or community-owned land.

This means that leopards must navigate their way across land dedicated to human development, agriculture or mining practices. As a result, they are exposed to an array of physiological, environmental and psycho-social factors that could cause stress.

Acute stress is essential for vertebrate survival. For example, hunting an impala may be stressful in the short term, but a successful kill equates to survival. In contrast, successive or simultaneous stressors experienced over prolonged periods of time, such as constantly having to avoid human interaction, can result in chronic stress. This, in combination with other factors could affect this already vulnerable species' long-term health and survival.

But how do you measure the stress levels within a leopard population without causing further distress? I set out to develop a method that would allow us to make a non-invasive assessment of stress levels in free-ranging leopards. It proved to be a useful approach.

My results indicate that although animals were relatively habituated at both sites, those living on the housing estate were more stressed than those in the game reserve. Pregnant females or those rearing cubs had the highest (617% higher) stress hormone levels of all the cats monitored. Overall, we found that wild male leopards showed less variation in their stress levels than females, regardless of whether they were in a protected area or not.

This method offers a new way for leopard biologists to monitor this elusive and iconic species. It can also inform the development of strategies to protect and conserve them.

Stress hormones

When we - leopards or humans - perceive a stressor, the central nervous system activates the release of hormones which act on the brain. Almost immediately, the pituitary gland releases hormones into the bloodstream and causes an almost instantaneous secretion of adrenalin. This mobilises energy which increases the heart rate and blood flow to the muscles so we have the physical means to confront the threat - or run away.

Over the next few hours, the adrenal glands release glucocorticoids - a type of steroid hormone - into the blood. These glucocorticoids (cortisol or corticosterone, depending on the species) are metabolised in the liver. After metabolism, they are then excreted via the bile into the gut and out of the body in the faeces. They can also travel via the kidneys to the bladder, to be excreted in the urine.

Previous studies have found that glucocorticoid concentrations are reliable indicators of disturbance experienced by an individual. That makes glucocorticoid metabolites very useful physiological indicators to measure stress. In this study we used scat to monitor the stress levels of free-ranging leopards.

We monitored two leopard populations. One consisted of seven known individuals living on a housing estate in Hoedspruit, a town located to the west of the Kruger National Park, South Africa's largest wildlife reserve. The other consisted of about 27 leopards living in a protected area adjoining the park.

Applying the science

We began the study by gathering faecal samples and observational data from leopards in two captive facilities. We used the faecal material to evaluate which of five chosen enzymeimmunoassays were best suited to pick up changes in the glucocorticoid concentrations in the faeces. Enzymeimmunoassays are widely accepted analytical tools for detecting particular antigens or antibodies in biological samples.

The captive leopards were monitored to determine how long food took to move through their systems, so we knew how long we needed to wait before getting a sample. It also enabled us to determine how long after defecation the hormones remained stable enough for measuring. We then used this information to compare the glucocorticoid concentrations in the faeces of our two groups of wild leopards.

Now that the method has been validated, we hope to use it to further examine how pregnancy, persecution outside of protected areas, levels of tourist activity and environmental factors contribute to the stress levels of this iconic African species.