Automated multi-day tracking of marked mice for the analysis of social behaviour

• A fully automated system to track multiple animals in a large arena without losing their identities is presented.
• The system learns unique bleach patterns on the mice's fur and tracks them during both dark and light cycles.
• Identification of six mice in the experimental setup was 97% correct during non-sleep intervals.
• As a proof of principle, we tracked groups of four mice and report social trends that develop across hours and days.

A quantitative description of animal social behaviour is informative for behavioural biologists and clinicians developing drugs to treat social disorders. Social interaction in a group of animals has been difficult to measure because behaviour develops over long periods of time and requires tedious manual scoring, which is subjective and often non-reproducible. Computer-vision systems with the ability to measure complex social behaviour automatically would have a transformative impact on biology. Here, we present a method for tracking group-housed mice individually as they freely interact over multiple days. Each mouse is bleach-marked with a unique fur pattern. The patterns are automatically learned by the tracking software and used to infer identities. Trajectories are analysed to measure behaviour as it develops over days, beyond the range of acute experiments. We demonstrate how our system may be used to study the development of place preferences, associations and social relationships by tracking four mice continuously for five days. Our system enables accurate and reproducible characterisation of wild-type mouse social behaviour and paves the way for high-throughput long-term observation of the effects of genetic, pharmacological and environmental manipulations.