UC Davis

Suboptimal fish stocking densities in experimental systems may elicit stress responses that can affect experimental results. Fish species, age and size, water chemistry and flow, and physical characteristics of the experimental system (e.g., tank, cage) are among the parameters to be considered when determining stocking densities. However, systematic studies to define fish densities minimizing stress in experimental systems are rarely performed. This is particularly true when working with species of low aquaculture value or a non-model test species such as the Delta Smelt (. Hypomesus transpacificus). The aim of this study was to use physiological stress biomarkers to determine suitable fish densities for specific experimental vessels routinely used for this species. We maintained late larval Delta Smelt (60. days post-hatch; dph) over a period of 24. h, at five different densities: 7, 14, 28, 42, and 56 fish per 8. L circular fish tank. We assessed whole body cortisol and transcriptomic biomarkers that lead to cortisol production to quantify stress levels. Both marker types delivered similar results. Cortisol levels were lowest at densities of 28 and 42 fish per tank, whereas lowest fish densities (7 and 14 fish per tank) evoked the highest stress levels. Genes such as Mineralocorticoid Receptor 1 and Glucocorticoid Receptor 2, as well as 11-Beta-Hydroxysteroid-Dehydrogenase-2 depicted the lowest expression levels at stocking densities 28 and 42, and elevated expression levels for stocking densities 7 and 14. Our data support the observations that late larval Delta Smelt should be exposed, acclimated, and cultured in groups rather than as individuals or in low numbers. This study indicates the importance of adequately defining experimental conditions that minimize stress, specifically when stress is measured as an endpoint. In addition to classical cortisol measurements, responses of the transcriptome also appear suitable in assessing stress responses in fish, and in determining optimal holding conditions, particularly if short-term responses are the study focus.Statement of relevance. The study highlights the importance of evaluating stress in order to determine species-specific stocking densities. The results are thus relevant to a wide audience in the field of aquaculture and experimental biology.