- Email: [email protected]
Local induction of IgT responses to pathogens and microbiota in the gill of rainbow trout
Abstracts / Fish & Shellfish Immunology 53 (2016) 58e93
O-037. Local induction of IgT responses to pathogens and microbiota in the gill of rainbow trout mez 1, Zhen Xu 1, 2, David Parra 3, Fumio Takizawa 1, Daniela Go Louise von Gersdorff Jørgensen 4, Scott LaPatra 5, J. Oriol Sunyer 1, * 1 Department of Pathobiology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA, USA 2 Department of Aquatic Animal Medicine, College of Fisheries, Huazhong Agricultural University, Wuhan, Hubei, 430070, China 3 Department of Cell Biology, Physiology and Immunology, Universitat noma de Barcelona, Barcelona, Spain Auto 4 Laboratory of Aquatic Pathobiology, Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiskberg DK-1870, Denmark 5 Research Division, Clear Springs Foods Inc., P O Box 712, Buhl, Idaho 83316, USA
Abstract Gas exchange structures are critical for acquiring oxygen, but they also represent portals for pathogen entry. Local mucosal immunoglobulin (Ig) responses against pathogens in specialized respiratory organs have only been described in tetrapods. We have previously shown that IgT is an Ig specialized in gut and skin mucosal immunity. Thus, we tested the hypothesis that IgT might play a pivotal role in mucosal immunity of teleost gills. In this study, we provide the first structural and functional characterization of all teleost Igs, including secreted IgD, IgM and IgT at a mucosal surface of a teleost fish. Here we show that IgTþ B cells represent the major B cell subset in the gill filaments. In contrast to reported results by others, we found that all gill B cells expressing surface IgM, also expressed surface IgD and that the percentage of B cells solely expressing either IgD or IgM was negligible. We found that the majority of bacterial microbiota in the gill mucosa is coated with IgT and, to a much lesser degree with IgM and IgD. More crucially, significant specific-IgT immune responses against Ichthyophthirius multifiliis (Ich) and Flavobacterium columnare were measured in the gill mucus, while pathogen-specific IgM responses were almost exclusively detected in the serum. Pathogen-specific IgD titers were absent both in gill mucus and serum. Importantly, we found significant IgTþ B-cell proliferative responses in the gill but not in the spleen or head kidney of fish that survived Ich infection. Moreover we also found that Ich- and F. columnare-specific IgT titers were locally produced by gill explants of survivor fish while they were absent in the spleen or head kidney explants of the same animals. In addition to showing that IgT is the main Ig player in gill mucosal immunity, the observed generation of local IgTþ B cell proliferative and pathogen-specific IgT responses in the gills provides the first demonstration of locally induced B cell and secretory Ig responses in the mucosa of a teleost. Moreover, this represents the first study in which a bacterial pathogen is shown to induce dominant IgT responses in a fish mucosal site, thus strongly suggesting that IgT is induced by a variety of pathogens in addition to parasites. Our findings also have special relevance from an applied perspective as they may lead to the development of fish vaccines and immunostimulants that have the capacity to induce gill IgT mucosal immune responses. Keywords: IgT, Ichthyophthirius multifiliis, Flavobacterium columnare, microbiota, gill * Corresponding author. Tel.: 215-573-8592; Fax number: 215-898-7887. E-mail address: [email protected] (J.O. Sunyer).
O-038. Leukocyte contributions to the induction and resolution of acute inflammation in teleost fish Dr. Daniel R. Barreda 1, 2, §, Jeffrey J. Havixbeck 1, Michael J. Trites 1, Michael E. Wong 1
71
1
Department of Biological Sciences, Canada Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB, T6G2P5, Canada 2
Acute inflammation drives early engagement of invading pathogens and contributes to the development of adaptive responses. This presentation will summarize recent experiments from our lab looking to define the contributions of leukocytes to the induction and resolution of acute inflammation in teleost fish. Using combinatorial cellular and molecular approaches we find that classic effectors such as macrophages and neutrophils elicit potent antimicrobial responses early in the acute inflammatory process. Interestingly, both leukocyte subsets also contribute to the resolution of inflammation. A timely shift in their phenotype, from proinflammatory to pro-resolving is largely driven by interactions with apoptotic cells at the site of inflammation. Examination of blood and hematopoietic compartments shows a tight correlation between hematopoietic blood cell production, their migration through circulation and discrete functional events at the infection site. Molecular effectors like transferrin also appear to play an important role in vivo, as previously suggested by in vitro experiments from other research groups. These molecules are also helping to better define the various phases of this acute inflammatory response. In all, we find that the capacity of leukocytes to polarize towards pro-inflammatory or resolution roles existed before the divergence of fish and tetrapods, over 450 million years ago. We also find unique features within the teleost fish group, such as their capacity to respond efficiently to pathogen challenge despite maintaining much lower numbers of circulating neutrophils (<5%). Keywords: acute inflammation, bony fish, leukocyte, immune response, evolution x Corresponding author. Tel.: 1(780)492 0375. E-mail address: [email protected] (D.R. Barreda).
O-039. Effects of IFNg administration on allograft rejection in ginbuna crucian carp Yasuhiro Shibasaki, Chihiro Hatanaka, Yuta Matsuura, Ryuichiro Miyazawa, Takeshi Yabu, Teruyuki Nakanishi§ Department of Veterinary Medicine, Nihon University, Fujisawa, Kanagawa 252-0880, Japan Abstract The rejection of allografts is primarily accomplished by cell-mediated immunity including T cells and IFNg in mammals. Recently, we identified four IFNg isoforms with antiviral activities in ginbuna crucian carp. In fish, however, the roles of IFNg isoforms in cell-mediated immunity remain unknown. Here we investigate the effects of administration of recombinant ginbuna IFNg (rgIFNg) isoforms in scale allograft rejection. IFNgrel 1 showed significantly higher expression at 5 days after allografting, while significant changes were not observed in other isoforms. Administration of rgIFNgrel 1 but not rgIFNgrel 2 nor rgIFNg 2 enhanced the allograft rejection. The number of CD4þ and CD8aþ cells in grafted scales and surrounding epidermis increased at an early stage of rejection, while that of sIgMþ cells increased at day 0 and day 5 in rgIFNgrel 1 administrated group. Accumulation of CD8aþ T cells from distal to medial epidermis of grafted scales was accelerated in rgIFNgrel 1 administrated group. Interestingly, mRNA expression of IFNg1 and IFNg2 was significantly upregulated after rgIFNgrel 1 administration, while that of IFNgrel 1 and IFNgrel 2 was not. Expression of granzyme was significantly higher at day 2 in rgIFNgrel 1 administrated group, while that of Perforin 1 and 3 was not. These results suggest that the administration of rgIFNgrel 1 induced the accumulation of T cell subsets at the allograft site and the secretion of effector molecules resulting in the promotion of allograft rejection. Present results also suggest the differential contribution of four IFNg isoforms to allograft rejection. Keywords: IFN gamma; allograft rejection; CD4; CD8; teleost
O-037. Local induction of IgT responses to pathogens and microbiota in the gill of rainbow trout mez 1, Zhen Xu 1, 2, David Parra 3, Fumio Takizawa 1, Daniela Go Louise von Gersdorff Jørgensen 4, Scott LaPatra 5, J. Oriol Sunyer 1, * 1 Department of Pathobiology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA, USA 2 Department of Aquatic Animal Medicine, College of Fisheries, Huazhong Agricultural University, Wuhan, Hubei, 430070, China 3 Department of Cell Biology, Physiology and Immunology, Universitat noma de Barcelona, Barcelona, Spain Auto 4 Laboratory of Aquatic Pathobiology, Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Frederiskberg DK-1870, Denmark 5 Research Division, Clear Springs Foods Inc., P O Box 712, Buhl, Idaho 83316, USA
Abstract Gas exchange structures are critical for acquiring oxygen, but they also represent portals for pathogen entry. Local mucosal immunoglobulin (Ig) responses against pathogens in specialized respiratory organs have only been described in tetrapods. We have previously shown that IgT is an Ig specialized in gut and skin mucosal immunity. Thus, we tested the hypothesis that IgT might play a pivotal role in mucosal immunity of teleost gills. In this study, we provide the first structural and functional characterization of all teleost Igs, including secreted IgD, IgM and IgT at a mucosal surface of a teleost fish. Here we show that IgTþ B cells represent the major B cell subset in the gill filaments. In contrast to reported results by others, we found that all gill B cells expressing surface IgM, also expressed surface IgD and that the percentage of B cells solely expressing either IgD or IgM was negligible. We found that the majority of bacterial microbiota in the gill mucosa is coated with IgT and, to a much lesser degree with IgM and IgD. More crucially, significant specific-IgT immune responses against Ichthyophthirius multifiliis (Ich) and Flavobacterium columnare were measured in the gill mucus, while pathogen-specific IgM responses were almost exclusively detected in the serum. Pathogen-specific IgD titers were absent both in gill mucus and serum. Importantly, we found significant IgTþ B-cell proliferative responses in the gill but not in the spleen or head kidney of fish that survived Ich infection. Moreover we also found that Ich- and F. columnare-specific IgT titers were locally produced by gill explants of survivor fish while they were absent in the spleen or head kidney explants of the same animals. In addition to showing that IgT is the main Ig player in gill mucosal immunity, the observed generation of local IgTþ B cell proliferative and pathogen-specific IgT responses in the gills provides the first demonstration of locally induced B cell and secretory Ig responses in the mucosa of a teleost. Moreover, this represents the first study in which a bacterial pathogen is shown to induce dominant IgT responses in a fish mucosal site, thus strongly suggesting that IgT is induced by a variety of pathogens in addition to parasites. Our findings also have special relevance from an applied perspective as they may lead to the development of fish vaccines and immunostimulants that have the capacity to induce gill IgT mucosal immune responses. Keywords: IgT, Ichthyophthirius multifiliis, Flavobacterium columnare, microbiota, gill * Corresponding author. Tel.: 215-573-8592; Fax number: 215-898-7887. E-mail address: [email protected] (J.O. Sunyer).
O-038. Leukocyte contributions to the induction and resolution of acute inflammation in teleost fish Dr. Daniel R. Barreda 1, 2, §, Jeffrey J. Havixbeck 1, Michael J. Trites 1, Michael E. Wong 1
71
1
Department of Biological Sciences, Canada Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB, T6G2P5, Canada 2
Acute inflammation drives early engagement of invading pathogens and contributes to the development of adaptive responses. This presentation will summarize recent experiments from our lab looking to define the contributions of leukocytes to the induction and resolution of acute inflammation in teleost fish. Using combinatorial cellular and molecular approaches we find that classic effectors such as macrophages and neutrophils elicit potent antimicrobial responses early in the acute inflammatory process. Interestingly, both leukocyte subsets also contribute to the resolution of inflammation. A timely shift in their phenotype, from proinflammatory to pro-resolving is largely driven by interactions with apoptotic cells at the site of inflammation. Examination of blood and hematopoietic compartments shows a tight correlation between hematopoietic blood cell production, their migration through circulation and discrete functional events at the infection site. Molecular effectors like transferrin also appear to play an important role in vivo, as previously suggested by in vitro experiments from other research groups. These molecules are also helping to better define the various phases of this acute inflammatory response. In all, we find that the capacity of leukocytes to polarize towards pro-inflammatory or resolution roles existed before the divergence of fish and tetrapods, over 450 million years ago. We also find unique features within the teleost fish group, such as their capacity to respond efficiently to pathogen challenge despite maintaining much lower numbers of circulating neutrophils (<5%). Keywords: acute inflammation, bony fish, leukocyte, immune response, evolution x Corresponding author. Tel.: 1(780)492 0375. E-mail address: [email protected] (D.R. Barreda).
O-039. Effects of IFNg administration on allograft rejection in ginbuna crucian carp Yasuhiro Shibasaki, Chihiro Hatanaka, Yuta Matsuura, Ryuichiro Miyazawa, Takeshi Yabu, Teruyuki Nakanishi§ Department of Veterinary Medicine, Nihon University, Fujisawa, Kanagawa 252-0880, Japan Abstract The rejection of allografts is primarily accomplished by cell-mediated immunity including T cells and IFNg in mammals. Recently, we identified four IFNg isoforms with antiviral activities in ginbuna crucian carp. In fish, however, the roles of IFNg isoforms in cell-mediated immunity remain unknown. Here we investigate the effects of administration of recombinant ginbuna IFNg (rgIFNg) isoforms in scale allograft rejection. IFNgrel 1 showed significantly higher expression at 5 days after allografting, while significant changes were not observed in other isoforms. Administration of rgIFNgrel 1 but not rgIFNgrel 2 nor rgIFNg 2 enhanced the allograft rejection. The number of CD4þ and CD8aþ cells in grafted scales and surrounding epidermis increased at an early stage of rejection, while that of sIgMþ cells increased at day 0 and day 5 in rgIFNgrel 1 administrated group. Accumulation of CD8aþ T cells from distal to medial epidermis of grafted scales was accelerated in rgIFNgrel 1 administrated group. Interestingly, mRNA expression of IFNg1 and IFNg2 was significantly upregulated after rgIFNgrel 1 administration, while that of IFNgrel 1 and IFNgrel 2 was not. Expression of granzyme was significantly higher at day 2 in rgIFNgrel 1 administrated group, while that of Perforin 1 and 3 was not. These results suggest that the administration of rgIFNgrel 1 induced the accumulation of T cell subsets at the allograft site and the secretion of effector molecules resulting in the promotion of allograft rejection. Present results also suggest the differential contribution of four IFNg isoforms to allograft rejection. Keywords: IFN gamma; allograft rejection; CD4; CD8; teleost