Prevailing role of mucosal immunoglobulins and B cells in teleost skin immune responses to bacterial infection

The skin of vertebrates is the outermost organ of the body and serves as the first line of defense against external aggressions. In contrast to mammals, the skin of early vertebrates like fish, lacks keratinization and has evolved to operate as a mucosal surface. As such, it has been shown to contain a skin-associated lymphoid tissue (SALT) in the epidermis. Prior to the discovery of IgT as a key mucosal immunoglobulin (Ig) in fish, several studies reported IgM-specific titers in the skin mucus upon pathogenic infection or vaccination. In contrast, a recent report showed IgT being the main Ig responding in the skin upon parasitic challenge. Whether IgT is the main Ig isotype elicited in the skin against bacterial pathogens, and whether such responses are locally induced within the SALT are important questions that remain unanswered. Here we analyzed the immune response of trout skin upon infection with one of the most widespread fish skin bacterial pathogens, Flavobacterium columnare (F. columnare). To gain insight into the pathogenesis and progression of the infection in skin, a recombinant F. columnare expressing green fluorescent protein (GFP) was constructed and showed to successfully invade the skin mucosa. This pathogen induced strong skin innate immune and inflammatory responses at the initial phases of infection. More critically, we found that the skin mucus of fish having survived the infection contained significant IgT- but not IgM- or IgD-specific titers against the bacteria. Moreover, we demonstrate the local proliferation and production of IgT+ B-cells and pathogen-specific IgT titers respectively within the SALT upon bacterial infection. Thus, our findings represent the first demonstration that IgT is the main Ig isotype induced by the skin mucosa upon bacterial infection, and that because of the large surface of the skin, its SALT probably represents a prominent IgT inductive site in fish. Author summary In contrast to mammalian skin, that of teleost fish is the largest mucosal surface containing a skin-associated lymphoid tissue (SALT). Thus far, IgT representing the prevalent immunoglobulin (Ig) in SALT have only been reported upon infection with a parasite. However, very little is known about the types of B cells and Igs responding to bacterial infection in the teleost skin mucosa, as well as the inductive or effector role of the SALT in such responses. To address these questions, we infected rainbow trout (Oncorhynchus mykiss) with F. columnare an important skin pathogen that causes significant losses both in wild and farmed fish populations. Remarkably, strong pathogen-specific IgT responses and accumulation of IgT+ B cells were induced in the cutaneous mucus and skin epidermis respectively upon infection. Evidence for the skin being an IgT inductive site was shown by the finding of local IgT+ B cells proliferation and pathogen-specific IgT tites in cultured trout skin explants. Thus, our findings demonstrate that IgT is the prevailing Ig isotype induced by the skin mucosa upon bacterial infection, and that the skin represents an IgT inductive site in fish.