Image Analysis Criteria for the Macroscopic Assessment of Skin Healing in Atlantic Salmon ^†

Abstract

Introduction: Fish skin is the first line of defense against the aquatic environment, acting as a physical, chemical, and immunological barrier. In addition to preventing pathogen entry, the skin and its mucus contribute to osmoregulation, innate immunity, and redox balance. Skin lesions—caused by mechanical damage, parasites, environmental stress, or handling—disrupt this barrier, increasing susceptibility to infections, inflammation, and production losses. Thus, efficient skin regeneration is essential for fish welfare and performance. Nutrition plays a key role in this process by providing substrates for epithelial repair, immune function, and antioxidant defense. Among dietary factors, zinc (Zn) is particularly important due to its involvement in cell proliferation, enzymatic activity, and maintenance of skin integrity. Objective: Our objective is to assess the effectiveness of image-based analysis in quantifying the skin healing process in Atlantic salmon fed diets supplemented with zinc. Methodology: The trial comprised three dietary treatments: a control diet with 42 mg Zn per kg (D1), and two diets supplemented up to 120 mg/kg of zinc, derived from inorganic (D2) or organic (D3) forms. Pit-tagged fish with an initial body weight (78 ± 0.1 g) were fed the diets for 75 days. After 15 days of experimental feeding, a standardized wound lesion (2.5 mm diameter × 0.5 mm depth) was inflicted in deeply anesthetized fish, with a disposable biopsy punch, in the dorsal area. After wound infliction, the fish resumed their normal feeding regime for the rest of the trial days. The progression of skin wound healing was assessed using standardized digital image analysis. High-resolution photographs of individual wounds were collected 8, 16, 24 and 32 days post-wounding. All images were acquired under standardized conditions with the inclusion of ArUco identifiers to enable a subsequent computer-assisted comparison. Morphometric parameters (wound width, diameter, perimeter and area) were used to assess wound contraction and closure over time. In parallel, a semi-quantitative visual scoring system was applied to each wound image to capture qualitative aspects of healing that are not fully described by morphometric data alone. Results: Full data analysis is currently underway, but the first results show beneficial effects of dietary zinc supplementation on the skin regenerative process. Conclusions: The combined use of objective digital measurements and standardized visual scoring enabled a comprehensive evaluation of wound healing progress, bridging quantitative tissue remodeling with biologically relevant phenotypic outcomes. This image-based framework provides a sensitive and reproducible approach for assessing dietary interventions targeting skin regeneration and barrier restoration in Atlantic salmon.