Introduction
Fish skin functions as the primary barrier against pathogens, mechanical damage, and environmental stressors, playing a crucial role in physiological homeostasis via mucus secretion and osmoregulation. Under intensive aquaculture practices, factors such as high stocking density, frequent handling, and temperature fluctuations often cause skin injuries, compromising the integrity of this protective barrier and increasing susceptibility to infections. Enhancing skin wound healing is thus critical for fish health and aquaculture productivity.
Zebrafish (Danio rerio ) have become a widely recognized model for studying vertebrate skin regeneration due to their rapid wound closure ability. Recently, a new laser ablation method has provided precise and reproducible control over wound size and depth, further enhancing experimental consistency. The present study utilized the laser ablation model in adult zebrafish to evaluate the efficacy of phytobiotic-based dietary additives in accelerating wound healing and promoting efficient skin regeneration.
Material and methods
Adult wild-type zebrafish were maintained at 28 °C in 10 L tanks with a 14-hour light/10-hour dark cycle and fed twice daily. Fish were randomly allocated into experimental groups (n=10 per group), receiving either phytobiotic-based dietary additives or PBS control by oral gavage at 500 mg/kg, administered twice weekly for 14 days before injury. Full-thickness skin wounds (~2 mm diameter) were induced using a 3 W laser on the left flank under anesthesia. Following laser ablation, fish were housed individually in aerated 3 L tanks with water renewal every two days, and dietary treatments continued until the trial conclusion. Wound areas were photographed at days 5, 10, 15, 20, 25, 30, and 35 post-ablation and analyzed using ImageJ software, expressed as the wound-closure percentage (WC%). At 10 days post-injury, skin samples were collected, embedded in paraffin, sectioned, and subjected to hematoxylin-eosin (H&E) staining and cytokeratin 18 (CK18) immunostaining to assess re-epithelialization and tissue remodeling activity.
Results
Following laser ablation, mortality rates remained low across all treatments and control groups, confirming minimal stress induced by laser ablation. Among the tested additives, APEX (Adisseo) , a phytobiotic-based formulation containing a mixture of medium-chain fatty acids and essential oils , significantly accelerated wound healing. Fish treated with the additive achieved 25% wound closure by 9.3 days and 50% closure by 16.2 days, substantially faster than the control group (26.7 days), representing nearly a 40% improvement in skin recovery. Histological assessments and CK18 immunostaining further confirmed enhanced re-epithelialization and tissue remodeling in APEX-treated fish. These findings indicate that dietary phytobiotic effectively stimulate both early and late phases of wound repair. Overall, the finding demonstrates strong potential as a dietary strategy to improve skin resilience and accelerate regeneration processes in fish, potentially reducing dependence on chemical treatments.