The newly developed method allows researchers to quickly and accurately measure stress hormones in snow leopards without the need for bulky equipment or specialized knowledge. It uses widely available equipment that can be transported to the field, allowing extraction of hormones from fecal samples and analysis on site.
This differs from existing approaches to hormone surveillance in wild animals, where fecal samples must be taken in laboratories for extraction and analysis of hormones. These approaches are particularly restrictive in remote regions, such as the Himalayas.
“Because conventional methods of hormone monitoring require frozen and refrigerated chemical reagents and laboratory equipment, it is almost impossible to use them on site. Explained Dr. Kodzue Kinoshita of Kyoto University and author of the study.
The new method developed by Dr Kinoshita extracts hormones from snow leopard droppings by hand-shaking a container containing the sample mixed with ethanol. A process called immunochromatography, used in pregnancy tests, detects hormone levels using test strips, and a smartphone app then analyzes them.
The accuracy of the new method was tested on fecal samples from captive snow leopards at Kohu Yuki Zoo, Asahiyama Zoo in Asahikawa City, and Nagoya Higashiyama Zoo and Botanical Gardens in Japan. Comparisons with conventional methods revealed that concentrations of similar hormones were extracted and that changes in these concentrations were also detected with precision.
Stress in animals can be associated with reduced reproductive function, so it is important for conservation to find out what in the environment is stressing the animals.
“Simple methods like this will allow researchers, rangers and zookeepers to quickly and easily assess the stress state of snow leopards.” Dr Kinoshita said. “Obtaining this information will be useful for animal welfare management and conservation planning.”
Dr Kinoshita added that the new technique could also be applied to captive snow leopards and other animals. “The hormone analyzed in this method is generally not species specific, so this method can be used to assess a variety of species, including domestic, experimental, zoo, and wild animals.”
Snow leopards are listed as “vulnerable” on the IUCN Red List and are threatened by poaching, reprisals for livestock attacks and climate change. It is feared that these continued pressures may increase stress in snow leopards, further contributing to the population decline.
Snow leopards are also notoriously difficult to study, the mountainous regions of Central Asia that they inhabit make it difficult to find big cats and limit researchers’ access to laboratories.
The new method developed by Dr. Kinoshita in this study involves adding ethanol to the collected droppings of snow leopards and then shaking the container with two zirconia beads in hand for two minutes to extract the hormones. The extract is then deposited on the immunochromatography test strip which turns red following an antigen-antibody reaction, indicating the presence of adrenal cortex stress hormones. A smartphone application is used to measure the concentration of hormones from the intensity of the color.
While this new method can be applied to other species, Dr Kinoshita warns that it will need to be tested for each species to ensure that the stress hormones detected in the faeces accurately reflect the stress in the animal, because these levels will vary from species to species.
“In a next step, I would like to apply this method to various other animals and make it more reliable.” Dr Kinoshita said. “I would also like to apply it not only to wild animals, but also to zoo animals and domestic animals to clarify the stress of these animals and improve their living environment.”
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