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Maternal immunity plays a vital role in protecting offspring from pathogen invasion, and it is present in nearly all vertebrates. As the most ancient vertebrates, teleost fish continuously secrete a substantial amount of mucus due to their unique living environment, and often exhibit schooling behavior. Previous studies have highlighted the presence of numerous immune components in the mucus of fish, that plays a crucial role in resisting pathogens. We hypothesize that this represents a distinctive maternal immune response mechanism in fish. In this study, we established a dextran sulfate sodium (DSS)-induced injury model by immersing zebrafish larvae in 0.5% DSS. This elicited robust inflammatory responses and showed DSS-induced alterations in microbial abundance via 16S rRNA sequencing. These changes included the disruption of normal gut structure; an increase in goblet cells; increased expression of inflammatory cytokines; and infiltration of neutrophils and macrophages into the gut, as well as an increase in pathogenic bacteria and a decline in probiotic bacteria. Furthermore, we conducted a mucus protection test by adding adult zebrafish mucus in fish tank water containing 0.5% DSS to test our hypothesis. As suspected, the mucus from adult zebrafish exhibited a protective role in defending zebrafish larvae against DSS-induced enteritis by alleviating excessive inflammatory responses and increasing probiotic abundance. In conclusion, our results confirm a significant role of adult fish mucus in the immune response of teleost fish to DSS-induced inflammation. Collectively, our findings show that the mucus of adult zebrafish may represent a novel form of maternal immunization, playing roles analogous to mammalian milk in immune regulatory functions.

The gallbladder (GB) microbiota plays critical roles in mammalian metabolism and immune homeostasis, and its relationship with human disease has been extensively studied over the past decade. However, very little is known about the interplay between GB microbiota and the immune functions of teleost fish, the earliest bony vertebrate with a GB. Therefore, this study sought to investigate the composition of the teleost GB microbiota and the potential mechanisms through which it affects mucosal immunity. In our results, we found that the GB mucosa (GM) and bile bacterial community shared a similar microbiological composition with that of the gut mucosa in naïve individuals. IHNV infection induced a profound GB inflammation and disrupted their microbial homeostasis followed by a strong anti-bacterial response. Interestingly, beneficial bacteria from the Lactobacillales order showed a significant increase in the abundance of the bile microbial community, whereas the structure of the Mycoplasmatales order in the gut microbial community was markedly changed. All in all, our study characterized the structure of the GB microbial ecosystem in teleost fish, and the fish GB microbiome shared a high similarity with the gut microbiota. More importantly, our findings offer solid evidence that the teleost GB evolved immune functions to preserve its mucosal microbial homeostasis, suggesting that both the microbiota and mucosal immunity of the GB might have co-evolved in early vertebrates.

The crosstalk between the immune system and microbiota drives an amazingly complex mutualistic symbiosis. In mammals, the upper respiratory tract acts as a gateway for pathogen invasion, and the dynamic interaction between microbiota and mucosal immunity on its surface can effectively prevent disease development. However, the relationship between virus-mediated mucosal immune responses and microbes in lower vertebrates remains uncharacterized. In this study, we successfully constructed an infection model by intraperitoneally injecting common carp (Cyprinus carpio) with spring viremia of carp virus (SVCV). In addition to the detection of the SVCV in the nose and pharynx of common carp, we also identified obvious histopathological changes following viral infection. Moreover, numerous immune-related genes were significantly upregulated in the nose and pharynx at the peak of SVCV infection, after which the expression levels decreased to levels similar to those of the control group. Transcriptome sequencing results revealed that pathways associated with bacterial infection in the Toll-like receptor pathway and the Nod-like receptor pathway were activated in addition to the virus-related Rig-I-like receptor pathway after SVCV infection, suggesting that viral infection may be followed by opportunistic bacterial infection in these mucosal tissues. Using 16S rRNA gene sequencing, we further identified an upward trend in pathogenic bacteria on the mucosal surface of the nose and pharynx 4 days after SVCV infection, after which these tissues eventually reached new homeostasis. Taken together, our results suggest that the dynamic interaction between mucosal immunity and microbiota promotes the host to a new ecological state.

Summary: The buccal mucosa (BM) is a critical first line of defense in terrestrial animals. To gain further insights into the evolutionary origins and primordial roles of BM in teleosts here we show that rainbow trout, a teleost fish, contains a diffuse mucosal associated lymphoid tissue (MALT) within its buccal cavity. Upon parasite infection, a fish immunoglobulin specialized in mucosal immunity (sIgT) was induced to a high degree, and parasite-specific sIgT responses were mainly detected in the buccal mucus. Moreover, we show that the trout buccal microbiota is prevalently coated with sIgT. Overall our findings revealed that the MALT is present in the BM of a non-tetrapod species. As fish IgT and mucus-producing cells are evolutionarily unrelated to mammalian IgA and salivary glands, respectively, our findings indicate that mucosal immune responses in the BM of teleost fish and tetrapods evolved through a process of convergent evolution. : Biological Sciences; Immunology; Omics; Transcriptomics Subject Areas: Biological Sciences, Immunology, Omics, Transcriptomics

The digestive tract is a unique series of organs that is inhabited by a range of commensal microbes while also exposed to an overwhelming load of dietary antigens. It is widely known that mammals have evolved complex and efficient immune strategies to protect the mucosa of the digestive tract. However, in the early vertebrates, the roles of mucosal immune defense and microbial communities in the different segments of the digestive tract are not well-understood. Here, we constructed a bath infection model with infectious hematopoietic necrosis virus (IHNV) in rainbow trout (Oncorhynchus mykiss). Importantly, following viral infection, we found that the IHNV distribution and the reactions of immune-related genes had similar trends that decreased across the digestive tract. Hematoxylin and eosin (H & E) and alcian blue (A & B) staining of the trout digestive tract showed that the pathological changes only occurred in the buccal and pharyngeal mucosal tissues. Moreover, the increased diversity of the microbial community was only detected in the buccal mucosa through 16S rRNA gene sequencing, suggesting that the magnitude of the immune response and microbial community changes are related to the IHNV load and the original microbial diversity. In addition, the loss of digestive tract dominant species and increased colonization of opportunistic bacteria were discovered in the buccal mucosal surface indicating that a secondary bacterial infection occurred in this mucosal tissue.

The olfactory organ of vertebrates receives chemical cues present in the air or water and, at the same time, they are exposed to invading pathogens. Nasal-associated lymphoid tissue (NALT), which serves as a mucosal inductive site for humoral immune responses against antigen stimulation in mammals, is present also in teleosts. IgT in teleosts is responsible for similar functions to those carried out by IgA in mammals. Moreover, teleost NALT is known to contain B-cells and teleost nasal mucus contains immunoglobulins (Igs). Yet, whether nasal B cells and Igs respond to infection remains unknown. We hypothesized that water-borne parasites can invade the nasal cavity of fish and elicit local specific immune responses. To address this hypothesis, we developed a model of bath infection with the Ichthyophthirius multifiliis (Ich) parasite in rainbow trout, Oncorhynchus mykiss, an ancient bony fish, and investigated the nasal adaptive immune response against this parasite. Critically, we found that Ich parasites in water could reach the nasal cavity and successfully invade the nasal mucosa. Moreover, strong parasite-specific IgT responses were detected in the nasal mucus, and the accumulation of IgT+ B-cells was noted in the nasal epidermis after Ich infection. Strikingly, local IgT+ B-cell proliferation and parasite-specific IgT generation were found in the trout olfactory organ, providing new evidence that nasal-specific immune responses were induced locally by a parasitic challenge. Overall, our findings suggest that nasal mucosal adaptive immune responses are similar to those reported in other fish mucosal sites and that an antibody system with a dedicated mucosal Ig performs evolutionary conserved functions across vertebrate mucosal surfaces.

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 mammalian skin, that of teleost fish lacks keratinization and has evolved to operate as a mucosal surface containing a skin-associated lymphoid tissue (SALT). Thus far, IgT representing the prevalent 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, in this study, we analyzed the immune response of trout skin upon infection with one of the most widespread fish skin bacterial pathogens, Flavobacterium columnare. 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 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. [ABSTRACT FROM AUTHOR]

The pharyngeal organ is located at the crossroad of the respiratory and digestive tracts in vertebrate, and it is continuously challenged by varying Ags during breathing and feeding. In mammals, the pharyngeal mucosa (PM) is a critical first line of defense. However, the evolutionary origins and ancient roles of immune defense and microbiota homeostasis of PM are still unknown. In this study, to our knowledge, we are the first to find that diffuse MALT is present in PM of rainbow trout, an early vertebrate. Importantly, following parasitic infection, we detect that strong parasite-specific mucosal IgT and dominant proliferation of IgT+ B cell immune responses occurs in trout PM, providing, to our knowledge, the first demonstration of local mucosal Ig responses against pathogens in pharyngeal organ of a nonmammal species. Moreover, we show that the trout PM microbiota is prevalently coated with secretory IgT and, to a much lesser degree, by IgM and IgD, suggesting the key role of mucosal Igs in the immune exclusion of teleost pharyngeal bacteria. Overall, to our knowledge, our findings provide the first evidence that pharyngeal mucosal immunity appear earlier than tetrapods. [ABSTRACT FROM AUTHOR]