Development of an 11-oxoetiocholanolone mini-kit for the quantification of faecal glucocorticoid metabolites in various wildlife species

As part of its mission to advance the field of wildlife endocrinology, the International Society of Wildlife Endocrinology aims to develop cost-effective antibodies and enzyme immunoassay kits that support research across a diverse range of species and sample matrices. To provide additional options for the quantification of faecal glucocorticoid metabolites (fGCMs), an antibody against 11-oxoetiocholanolone-17-carboxymethyl oxime (CMO) was generated in rabbits, and an enzyme immunoassay incorporating a horseradish peroxidase-conjugated label and 11-oxoetiocholanolone standard has been developed, designed for use with anti-rabbit IgG secondary antibody coated plates. This mini-kit was used to quantify glucocorticoid metabolites with a 5 beta-3 alpha-ol-11-one structure in faecal extracts from 23 species: African and Asian elephants, Alpine chamois, American bison, Bengal tiger, blue wildebeest, blue-and-yellow macaw, brushtail possum, cape buffalo, fat-tailed dunnart, Florida manatee, ghost bat, giraffe, golden langur, Gould's wattled bat, hippopotamus, Leadbeater's possum, mandrill, okapi, roan antelope, samango monkey, short-beaked echidna, and western lowland gorilla. Pharmacological (adrenocorticotropic hormone challenge) and biological (inter-zoo translocation, wild capture, social disruption, illness/injury and veterinary intervention) challenges resulted in expected increases in fGCM concentrations, and in a subset of species, closely paralleled results from a previously established immunoassay against 11-oxoetiocholanolone-17-CMO. Two additional species tested, Krefft's glider, which showed contradictory results on this assay compared to a previously validated enzyme immunoassay (EIA) and Ankole cow, where the magnitude increase post-event did not quite reach the 2-fold change criteria, highlight that differences in excreted faecal metabolites across species mean that no EIA will be suitable for all species. This assay provides a valuable new option for assessing adrenal activity across taxa using a group-specific antibody. Future studies should put similar emphasis on validation to determine optimal assay choice for measuring fGCMs in a variety of species. Glucocorticoid hormones (cortisol, corticosterone) are useful indicators of the stress response, but for many wildlife species, non-invasive approaches and appropriate tools for measurement of excreted metabolites are required. Here, we report the development and validation of a group-specific enzyme immunoassay to quantify glucocorticoid metabolites in the faeces of 23 species.

South East Zoo Alliance Reprod & Conservat
Chester Zoo
Disney´s Animal Kingdom, Animal Programs Administration
Arbor Assays Inc
Smithsonian Institution
University of Queensland
La Trobe University
University of Pretoria