Shkurov AY, Ostras DA, Harkavenko VV. 2026. Effects of
caffeine exposure on the behavior of the invasive marbled crayfish Procambarus
virginalis. In: Pilay AP (Editor-in-chief), VetBioConnect: Young Scientists
Conference, 3-4 June 2026, Kharkiv, pp. 91-93.
https://doi.org/10.36016/VBC-2026 (Abstract only.)
Background. Caffeine is a widespread pharmaceutically active
compound that enters surface waters mainly with municipal wastewater and may
alter the behavior of aquatic organisms even at sublethal concentrations.
Behavioral endpoints are especially important in ecotoxicology because changes
in movement, shelter use, social interactions, and aggression can affect
survival, dispersal, and population-level responses before mortality occurs.
Crayfish are suitable model organisms for such studies because they demonstrate
distinct spatial, social, and agonistic behavior that can be recorded and
quantified under laboratory conditions. The marbled crayfish Procambarus
virginalis is of particular interest because it is a parthenogenetic, highly plastic,
and invasive species capable of rapidly establishing new populations from aquarium
releases. For invasive hydrobionts, pollutant-induced behavioral changes may
influence both individual stress tolerance and the ecological success of populations
in transformed freshwater ecosystems. Previous studies have shown that psychoactive
pollutants can modify locomotor activity, shelter-related behavior, and risk-taking
in aquatic invertebrates, including crayfish. However, the behavioral effects
of caffeine on P. virginalis remain insufficiently described. Therefore, short-
term laboratory exposure combined with ethogram-based video analysis can
provide useful preliminary data on how this invasive crayfish responds to
caffeine contamination.
Aim. The aim of this study was to evaluate the effects of
caffeine exposure on the behavioral
structure of the invasive marbled crayfish Procambarus virginalis and to describe the temporal dynamics of changes
in locomotor, anxiety-related, shelter- related, and aggressive activity over
24 hours.
Materials and methods. The study was carried out under
laboratory conditions using video recording followed by ethogram analysis in
BORIS software. Marbled crayfish were kept in an open 20 L container. In the
control series, baseline behavior was recorded without the addition of any
substance. In the experimental series, behavior was recorded after a single
addition of caffeine at a dose of 200 mg per 20 L of water, corresponding to 10
mg/L. Behavior was assessed at 0, 6, 12, and 24 h after exposure. The duration
and frequency of the following behavioral categories were analyzed: Move, Rest,
Swim, Shelter, Wall, Field, Aggression, Attacking pose, and Ball of crayfish.
The control and experimental observations were compared to identify changes in
the distribution of behavioral forms and in the temporal structure of activity.
Results. In the control group, the behavioral profile
remained relatively stable throughout the observation period: Rest predominated
(60‒68% of the time), locomotor activity remained moderate (15‒22%), and
aggressive or anxiety-related reactions were rare. During the first 10 min
after caffeine addition, crayfish demonstrated pronounced hyperactivity and
stress-related orientation: Move increased to 33.5%, Wall to 12.5%, Ball of
crayfish to 18.9%, and Attacking pose to 18.7%. After 6 h, the behavioral
pattern changed sharply: Rest reached 86.3%, whereas Aggression increased to
25.0% and Attacking pose to 25.7%, indicating motor exhaustion combined with
persistent social tension. After 12 h, locomotor activity almost disappeared
(Move 0.6%), while Attacking pose remained high (24.8%), suggesting postural
tension under conditions of minimal mobility. After 24 h, Rest again
predominated (73.9%), Aggression decreased to 6.6%, Attacking pose decreased to
4.2%, and episodic shelter use appeared.
Conclusions. Caffeine exposure caused a phase restructuring
of the behavioral pattern of P. virginalis: from acute hyperactivity and
anxiety through aggressive exhaustion to a relatively stable state of
low-mobility adaptation. The results indicate that the behavior of marbled
crayfish may be used as a sensitive indicator of caffeine and other
pharmaceutical pollutants in aquatic environments. This approach can be applied
in further studies of behavioral ecotoxicology of invasive hydrobionts and in the
assessment of ecological risks in transformed freshwater ecosystems.
Keywords: marbled crayfish • Procambarus virginalis • caffeine • behavioral responses • invasive species • ecotoxicology.
