A paired longitudinal comparison of fecal and serum cortisol in Holstein transition dairy cows.

Transition dairy cows experience substantial physiological stress around calving. Serum cortisol reflects acute hypothalamic-pituitary-adrenal activity, whereas fecal cortisol metabolites (FCM) provide a non‑invasive measure of cumulative cortisol secretion. This study quantified the within‑cow association between log10-transformed FCM and serum cortisol, characterised temporal trajectories and lags, evaluated the ability of FCM to predict serum cortisol, assessed links with oxidative stress markers, and examined effect modification by the temperature-humidity index (THI). Twenty multiparous Holstein cows were sampled on nine days relative to calving. Linear mixed models (LMMs), generalised additive mixed models (GAMMs), cross‑correlation functions and cross‑validated predictive models were used. A significant positive association between FCM and serum cortisol was found, but prediction failed to meet prespecified calibration criteria, as the calibration slope (0.843 [95% CI 0.172-1.514]) fell outside the prespecified 0.90-1.10 range and the calibration intercept (0.356 [-1.085-1.799]) fell outside the - 0.05 to 0.05 range. Peak days differed across cows, and no systematic lag was detected; instead, we observed substantial cow‑level variability in lag structure. FCM was not associated with oxidative markers, and THI did not modify the serum-FCM relationship. These results suggest that FCM complements rather than substitutes for serum cortisol in transition cows.