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The Predictive Accuracy of Methods Commonly Used for Evaluating Animal Distress.

Emily Leitner¹, Annika Thämlitz¹, Muhammad Imran Khan¹

¹Rudolf-Zenker-Institute of Experimental Surgery, Rostock University Medical Center, Rostock, Germany.

FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology

|June 8, 2026

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Accurate animal distress assessment is crucial. Distress scoring and body weight changes are reliable indicators across various conditions, outperforming burrowing and nesting, especially in low-distress scenarios.

Area of Science:

Animal Welfare Science
Preclinical Research Methods
Veterinary Medicine

Background:

Accurate distress assessment is vital for animal welfare and scientific validity in research.
Current behavioral and physiological indicators' predictive value across disease models and severity grades is not well-defined.
Evaluating multiple indicators is necessary to establish reliable distress assessment protocols.

Purpose of the Study:

To evaluate the discriminatory performance of body weight changes, distress scoring, burrowing, and nesting in mice across different disease models and severity grades.
To assess the cross-project accuracy of these indicators when optimal cut-offs are applied to independent datasets.
To determine the reliability of these distress indicators under varying levels of distress.

Main Methods:

Keywords:

animal welfare bile duct ligation chronic pancreatitis distress receiver operating curves refinement robustness transmitter implantation

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Utilized ten datasets involving transmitter implantation, bile duct ligation (BDL), and chronic pancreatitis (CP) in mice.
Employed receiver operating characteristic (ROC) analyses to determine the area under the curve (AUC) for each indicator.
Applied optimal cut-offs derived from one dataset per model to independent datasets for cross-project accuracy assessment.

Main Results:

Under moderate to high distress (BDL, acute implantation), distress score (AUC ≥ 0.96) and body weight (AUC ≥ 0.76) showed excellent discriminative performance.
Under low distress (CP, recovery), discrimination was reduced: body weight (AUC ≥ 0.62), distress score, burrowing, and nesting (AUC ≥ 0.50).
Distress score and body weight maintained high accuracy (≥ 86%) on independent datasets under moderate to high distress, while burrowing and nesting were less reliable (≥ 65%).

Conclusions:

Distress score and body weight are reliable indicators of animal distress, particularly under moderate to high distress conditions.
Burrowing and nesting behaviors can supplement distress evaluation under moderate to high distress but are less reliable than body weight and distress score.
The reliability of all tested indicators, especially behavioral measures, significantly decreases under low-distress conditions, highlighting the need for careful interpretation.