Atlantic salmon is one of the most important aquaculture species worldwide, accounting for 4.5 % of global finfish trade, and demand is steadily increasing ( FAO 2020) . However, production is hindered by skin diseases such as sea lice and winter ulcerating bacterial infections, which caus e substantial welfare issues as well as economic losses (Fish Health report 2022) . W ith both diseases on the rise, there has been a growing interest on salmon skin biology. One of the main characteristics of skin is its ability to heal, enabled by the presence of specialized cells that work together to repair damage and restore the integrity of the tissue (Hou et al. 2020). Knowledge of the cellular composition and genetic response to infection, wounding and healing of Atlantic salmon skin is needed in order to gain a better understanding of how these pathogens are bypassing and suppressing the healing capabilities of skin.
Materials and Methods
Samples for single cell sequencing and spatial transcriptomics were collected from two healthy skin samples and four mechanically wounded skin samples at 2 and 14 days post wounding (two samples each time point), representing the early acute inflammatory phase and the tissue repair phase, respectively . Mechanical wounds were introduced by a sterile punch biopsy tool as described by (Sveen et al. 2018).Single-nuclei RNA-seq libraries were generated for all samples using a 10x Chromium and following standard 10x protocols. Spatial transcriptomics libraries where prepared using the 10x visium protocol. Library mapping was performed with STAR (Kaminow et al. 2021) with v3.1 of the genome of Atlantic salmon as reference. Results were analysed using Seurat (Stuart et al. 2019).
We generated single cell atlases across a wound healing time course. We identified pluripotent mesenchymal stem cells that differentiate into diverse cell types including adipose, bone and vascular cells and appear to be crucial in the wound healing process. We then used spatial transcriptomics techniques to locate these cell types in the tissue, and demonstrate their proximity to the wound and importance to the healing process (Figure 1). We further interrogate the role of these cells in the healing process, including their interactions with keratinocytes and immune cells
This work represents a major enhancement in our understanding of the wound healing process in Atlantic salmon, identifying and characterising the key role of pluripotent mesenchymal stem cells . Importantly, it provides a framework for comparison of healthy wound healing and healing of pathogen-afflicted wounds, such as those caused by sea lice and winter ulcerating bacteria .
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Fish health report (2022) https://www.vetinst.no/rapporter-og-publikasjoner/rapporter/2022/fish-health-report-2021
Sveen et al. (2018). High fish density delays wound healing in Atlantic salmon (Salmo salar). Sci Rep 8, 16907https://doi.org/10.1038/s41598-018-35002-5
Hou et al. (2020) Cellular diversity of the regenerating caudal fin. sci.avd. 33 DOI: 10.1126/sciadv.aba2084
Kaminow et al. (2021). STARsolo: accurate, fast and versatile mapping/quantification of single-cell and single-nucleus RNA-seq data. Biorxiv, 2021-05.
Stuart et al. (2019). Comprehensive integration of single-cell data. Cell, 177(7), 1888-1902.