Validation of gut transit rate assessment methodology and the mitigation of sampling stress in Atlantic salmon, Salmo salar

Gut transit rate is an important component of the digestive process in fish, particularly in the context of aquaculture, where factors such as rearing temperature and dietary formulation impact on-farm performance. As such, developing and validating research methodologies that explore these critical physiological responses in fish is vital in the context of precision aquaculture practices. However, there are inherent methodological constraints of gut transit rate assessments, particularly when assessing the effect of sampling on the stress response of fish. In light of this, the present study aimed to assess the gut transit rate and plasma cortisol levels of Atlantic salmon ( Salmo salar ) subjected to the more commonly applied, repeated sampling model that results in multiple tanks disturbances (e.g., 2, 4, 8, 16 and 24 h post-feed) in comparison to one requiring only a singular tank disturbance at one of the designated time points. An inert beaded marker was integrated into diet formulations to determine the transit rate of digesta in three major gastrointestinal tract regions: the stomach, mid-intestine, and distal intestine over a period of 48 h following the provision of food. A significantly higher plasma cortisol concentration was recorded in fish subjected to a repeated disturbance, particularly at 16 h post feed (22.99 ± 6.88 ng mL −1 ), compared to fish sampled from tanks subjected to a single sampling event (11.35 ± 0.92 ng mL −1 ). Despite this, gut transit rate was generally unaffected by sampling design over the duration of the study with both groups of fish displaying a similar rate of gut transit. This suggests that repeated tank sampling (i.e. sampling multiple fish from the same tank at multiple timepoints) is suitable to assess gut transit rate in fish as it does not significantly confound gut transit rate assessments. It is envisioned that this data will contribute to the growing understanding of the digestive process of Atlantic salmon and the relationship with stress response in fish and experimental design. Moreover, it should strengthen future studies aimed at assessing key digestive processes in cultured species. • Ballotini beads in feed were used to assess gut transit rate in Atlantic salmon. • Tanks were subjected to either repeated sampling or a single sampling event. • The effect of both designs on gut transit and plasma cortisol in fish was assessed. • Repeated tank sampling had minimal effect on the gut transit rate of fish. • Plasma cortisol levels in fish increased in tanks subjected to repeated sampling.