| As the outermost organ of the organism’s body,fish skin is continuously exposed to a wide array of potentially harmful insults in the aquatic environment.Meanwhile,the constant exposure has also necessitated the evolution of a complex and well-coordinated immune network in the skin,which ensures that adequate immune responses are mounted in response to antigenic challenge while maintaining overall homeostasis.In contrast to mammalian skin,teleost skin has been considered as mucosal surface which serves as the first line of defense against invading pathogens.Besides,teleost skin also contains a skin-associated lymphoid tissue(SALT)that elicits gut-like immune responses against antigen stimulation.Immunoglobulins(Igs)are the key components of adaptive immunity.Teleosts represent the most ancient bony vertebrates containing immunoglobulin(Ig)within the non-tetrapods.In contrast to mammals,as far as we know,only three Ig isotypes have been identified in teleosts(Ig M,IgT/Ig Z,and Ig D).Teleost IgT has been identified to play a specialized role in fish mucosal immune responses akin to that of Ig A in mammals.Remarkably,previous studies have shown parasite-specific IgT and Ig M titers in teleost skin mucus and serum,respectively,indicating a specialized role of IgT in skin mucosal immune responses.However,little is known about the B cells and Immunoglobulins(Igs)responds to bacterial or viral infection in teleost skin mucosal immune system.Whether bacterial and viral infection can also induce the specific responses of IgT an even the local responses of mucosal B cells in teleost skin are thus far unknown.Moreover,exposed to the water environment and the pathogens therein,teleost skin is also known to be colonized by diverse microbial communities.However,little is known about the interactions between microbiota and the teleost skin mucosal immune system,especially dynamic changes about the interactions under pathogen infection.Here,we’ve constructed infected models with rainbow trout(Oncorhynchus mykiss),which was experimentally exposed to Flavobacterium columnare,IHNV,and Ichthyophthirius multifiliis,respectively,since the bacteria and parasite mainly invade the skin of fish and cause lesions on the surfaces.Using the models,we’ve detected the morphological changes and immune responses including innate and adaptive response in trout skin following pathogenic infection.We mainly studied the functions of Igs and B cells in trout skin against different pathogens.Then we also detected the relations among pathogen,host immunity and commensals in trout skin.At last,we were surprised to find that IgT may take part a import role in the innate immune responses against Ich infection.1. Taken together,our results indicate that Flavobacterium columnare,IHNV,and Ichthyophthirius multifiliis,are all able to invade the skin mucosal tissue and cause significant histological responses.By hematoxylin&eosin(HE)and alcian blue(AB)staining,morphological changes were observed in both the thickness of the skin epidermis and the number of mucus cells in the experimental trout skin with paraffin sections.Our results revealed that bacterial and viral invasion destroyed the epidermal cells of fish skin,while parasitic invasion could increase the width of skin epidermis and the number of mucus cells.2. Our findings showed that trout skin is an ancient mucosal surface that elicits strong innate and adaptive immune responses to pathogenic infection.By qRT-PCR and RNA-Seq analysis,we identified SALT-related immune molecules that respond to F.columnare,including antibacterial peptides,pro-inflammatory cytokines,chemokines,complement factors,and Igs,all of which are known to be crucial in the teleost skin immune system.Similarly,IHNV could induced antiviral genes and also pro-inflammatory cytokines,chemokines,complement factors,and Igs in trout skin.While Ich infection could upregulate not only interleukins,chemokines,complement factors,lectin,and mucins,but also antimicrobial peptides.3. Teleost IgT and IgT~+B cells play the dominant role in skin mucosal immune responses to different kinds of pathogens including bacteria,virus,and parasite.Microbial as well as virus exposure can also elicit a dedicated mucosal immunoglobulin response the same as that of parasites in teleost skin.Strikingly increased IgT concentration and strong pathogen-specific IgT responses were detected in the cutaneous mucus.Accumulation of IgT~+B cells were also found in trout skin after pathogen infection.4. Our results demonstrate that IgT responses against the pathogen were mainly limited to the skin whereas Ig M responses were almost exclusively detected in the serum.In addition,SALT is both an inductive and effector immune site of B cells responses in teleost fish.5. Importantly,we showed for the first time the local production of IgT and proliferation of IgT~+B cells in the skin mucosa after bacterial challenge,thus providing the first example of locally induced mucosal Ig immune responses in the skin of a non-tetrapod species.6. Parasitic invasion can break the microbial homeostasis in trout skin.High diversity values of microbiota in trout skin from the experimental groups were discovered.7. Our results suggest for the first time that parasitic infection could inhibit symbionts and offer opportunities for other pathogens’secondary infection in teleost skin,which were mainly characterized by a decreased abundance of skin commensals(lose of Proteobacteria)and increased colonization of opportunistic bacteria(increased intensity of Flavobacteriaceae).8. Teleost IgT may take part in the early immune responses to parasite infection in the skin mucosa.Increased IgT and IgT~+B cells were detected in trout skin at early times following Ich infection.At 7 days post infection,Ich trophonts were localized within the skin epidermis and intensely stained with anti-IgT,but little with anti-Ig M antibodies... |
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