Little agreement among methodologies to determine fecal glucocorticoid metabolites in a mountain ungulate

Faecal glucocorticoid metabolites (FGMs) have gained relevance in ecological studies and population monitoring, allowing non-invasive remote sampling without the need to capture and handle animals. Enzyme immunoassays (EIAs) and radioimmunoassays (RIAs) are commonly used for FGM determination. Although these methods should be validated for each species, nonvalidated tests are still widely used to determine FGMs in wildlife. Near-infrared reflectance spectroscopy (NIRS) is a predictive method requiring calibration against reference methods used to assess FGMs in wildlife. EIAs and RIAs have been utilized to determine FGMs in chamois (Rupicapra spp.), a medium-sized mountain ungulate. This study aims to assess the potential of NIRS to determine FGMs and to evaluate the correlation among analytical methods used to determine FGMs in chamois. Faecal samples from 125 Alpine chamois (Rupicapra rupicapra rupicapra) and 125 Pyrenean chamois (R. pyrenaica pyrenaica) were collected from the field, frozen at - 20 C-degrees, lyophilized, grounded, and scanned using a NIRSystems 5000 monochromator over a 1108-2492 nm wavelength. After this non-destructive NIRS analysis, FGMs were extracted and analysed using four immunoassays previously used in chamois studies: a 125-I-corticosterone RIA, a cortisol EIA, and two 11-oxoetiocholanolone EIAs (72a and 72T). Only the 11-oxoetiocholanolone 72T EIA has been validated for Alpine chamois. NIRS predictions were calibrated and cross-validated for each of the four immunoassays. The correlation among the four immunoassays was assessed using Spearman's rank. The coefficient of determination for NIRS calibration (R2) values ranged from 0.37 to 0.75, and the ratio of performance to deviation values from 1.2 to 1.6. Therefore, NIRS could not predict FGM concentration in chamois faeces. This could be due to the complexity and variability of the FGM detected by the immunoassays as reference methods, and by potential interference of other compounds in the faecal matrix. The correlation among the immunoassays was low overall. As a conclusion, NIRS cannot be recommended for measuring FGMs in chamois. The low correlation among the immunoassays used for FGM determination raises concern about the reliability of previous studies using non-validated methods in chamois. Only biologically