Establishing a Percutaneous Infection Model Using Zebrafish and a Salmon Pathogen

SIMPLE SUMMARY: The epidermis and mucus layer of fish act as barriers that protect them against waterborne pathogens, and provide niches for symbiotic microorganisms that benefit the host’s health. However, our understanding of the relationship between fish skin bacterial flora and fish pathogen infection is limited. In order to elucidate this relationship, an experimental model for infection through fish skin is necessary. Such a model must also pose a low biohazard risk in a laboratory setting. We established a percutaneous infection model using zebrafish (Danio rerio), a typical fish experimental model, and Yersinia ruckeri, a salmon pathogen. Our experimental data indicate that Y. ruckeri colonizes niches on the skin surface generated by transient changes in the skin microflora caused by stress, dominates the skin bacterial flora, occupies the surface of the fish skin, invades the fish body through injury, and finally, causes fatal enteric redmouth disease. This percutaneous infection model can be used to study the interaction between fish skin bacterial flora and fish pathogens in water, or the relationship between pathogens and the host’s skin immune system. ABSTRACT: To uncover the relationship between skin bacterial flora and pathogen infection, we developed a percutaneous infection model using zebrafish and Yersinia ruckeri, a pathogen causing enteric redmouth disease in salmon and in trout. Pathogen challenge, either alone or together with pricking by a small needle, did not cause infection of the fish. However, cold stress given by water temperature shift from the optimum 28 °C for zebrafish to 20 °C caused fatal infection of injured fish following pathogen challenge. We investigated the effects of cold stress, injury, and pathogen challenge, alone and in combination, on fish skin bacterial flora using 16S rDNA metagenomics. We found that cold stress drastically altered the skin bacterial flora, which was dominated by Y. ruckeri on infected fish. In addition, fish whose intrinsic skin bacterial flora was disrupted by antibiotics had their skin occupied by Y. ruckeri following a challenge with this pathogen, although the fish survived without injury to create a route for invasion into the fish body. Our results suggest that the intrinsic skin bacterial flora of fish protects them from pathogen colonization, and that its disruption by stress allows pathogens to colonize and dominate their skin.