- Email: [email protected]
Characterization of antigen sampling cells in rainbow trout gill epithelium
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Abstracts / Fish & Shellfish Immunology 53 (2016) 58e93 1 Center of Evolutionary and Theoretical Immunology Department of Biology University of New Mexico, New Mexico, USA 2 Research Division Clear Springs Foods Inc, Buhl, Idaho, USA
Abstract Mucosal surfaces are multifunctional barriers that need to orchestrate a variety of physiological functions. One of the mechanisms by which mucosal surfaces carry immunological functions without compromising other physiological roles is the establishment of regional immunity. Regional immunity can be generated by selective expression of chemokines, chemokine receptors and adhesion molecules within mucosal microenvironments. In the olfactory organ of animals, immune responses may result in damage to the olfactory sensory neurons and, as a consequence, compromise olfaction. The olfactory organ of teleost fish is histologically divided into two distinct areas, a sensory or neuroepithelial region in the lateral regions and valleys if the lamellae and a non-sensory region located at the tip of each lamella characterized by presence of goblet cells. Thus, we hypothesized the presence of two unique immune microenvironments corresponding with these two regions in rainbow trout. In line with our hypothesis, we found clusters of CD8þ T cells at the mucosal regions of lamellae whereas few numbers were found in the neuroepithelium. Nasal CD8þ T cells displayed a mucosal phenotype similar to those found in the gut. Laser capture microdissected tip versus lateral samples of control trout olfactory tissue were collected and RTqPCR revealed a higher expression of the chemokine and chemokine receptor CCL19 and CCR7, the adhesion molecules L-selectin, ICAM, VCAM, and integrin b2 and several T cell markers in the mucosal tip compared to the neuroepithelium. Moreover, MHC-II immunostaining indicated that the mucosal tip preferentially participates in antigen presentation within the trout nasal mucosa. Finally, in response to nasal vaccination, cell proliferation occurs in the tip mucosal region and not the neuroepithelium. Our results support the idea of compartmentalized immune responses within the nasal mucosa of teleosts that likely evolved as a sparing mechanism to protect olfactory sensory function. Keywords: Regional immunity, NALT, trout, nasal immunity, mucosal immunity * Corresponding author. Tel.: þ1 505 277 0039. E-mail address: [email protected] (I. Salinas).
O-070. Characterisation and bioactivities of type I interferon subtype E and F in salmonids Thitiya Summathed 1, *, Jun Zou 1, Simon Wadsworth 2, Chris Secombes 1 1
Scottish Fish Immunology Research Centre, Institute of Biological and Environmental Sciences, University of Aberdeen, Tillydrone Avenue, Aberdeen, AB24 2TZ, UK 2 EWOS Innovation A.S., Dirdal, N-4335, Norway Abstract Salmonids are important commercial species in the European aquaculture industry. Many viral diseases can impact on the industry, leading to economic losses. Interferons (IFNs) are cytokines involved in antiviral defence, and study of their regulation and function may shed light on how fish can defend themselves against viral infection. Fish type I IFNs are classified into two groups based on cysteine residues in the mature peptide, namely group I and II IFNs, each of which is further divided into 3 subtypes in salmonids. The latest new subtypes, IFN-e and IFN-f, have only recently been identified and their biological activities have still to be characterised. In this presentation, IFN-e and ef genes in Atlantic salmon (Salmo salar) and their bioactivities were reviewed and compared to previous studies of other type I IFN subtypes (a-d). The type I IFN subtypes showed different expression patterns between the 2 groups in cell lines and primary head kidney cells stimulated with poly I:C and in heart and head kidney of cardiomyopathy syndrome (CMS) infected salmon. Transcript levels could be correlated to the antiviral response and the IFN-e and IFNef
recombinant proteins were shown to induce Mx gene expression which is known to mediate antiviral activity. This data together with the reactivity of specific antibodies to the subtypes provides a better understanding of the antiviral effects of type I IFNs in salmonids. Keywords: Salmonids, type I interferons, bioactivities * Corresponding author. E-mail address: [email protected] (T. Summathed).
O-071. CK9, an ancient rainbow trout chemokine that attracts and regulates B lymphocytes and macrophages Carolina Aquilino 1, Aitor G. Granja 1, Rosario Castro 1, Tiehui Wang 2, Beatriz Abos 1, David Parra 3, Christopher J. Secombes 2, Carolina Tafalla 1, § n en Sanidad Animal (CISA-INIA). Valdeolmos, Centro de Investigacio Madrid, Spain 2 Scottish Fish Immunology Research Centre, School of Biological Sciences, University of Aberdeen, Aberdeen AB24 2TZ, UK 3 Animal Physiology Unit, Department of Cell Biology, Physiology and Immunology, School of Biosciences, Universitat Autonoma de Barcelona, Cerdanyola del Valles, Spain 1
Abstract CK9 is a rainbow trout (Oncorhynchus mykiss) CC chemokine phylogenetically related to mammalian CCL25. Although CK9 is known to be transcriptionally regulated in response to inflammation particularly in mucosal tissues, its functionality has never been revealed. In the current work, we have demonstrated that CK9 has a strong chemotactic capacity for both B cells (IgMþ and IgTþ) and macrophages. Along with its chemotactic capacities, CK9 modulated the MHC II turnover of B lymphocytes and upregulated the phagocytic capacity of both IgMþ cells and macrophages. Although CK9 had no lymphoproliferative effects, it increased the survival of IgTþ lymphocytes. Furthermore, we have established that the chemoattractant capacity of CK9 is strongly increased after pre-incubation of leukocytes with a T-independent antigen, whereas B cell receptor (BCR) cross-linking strongly abrogated their capacity to migrate to CK9, indicating that CK9 preferentially attracts B cells at the steady state or under BCR-independent stimulation. These results point to CK9 being a key regulator of B lymphocyte mucosal trafficking in teleost fish, able to modulate innate functions of teleost B lymphocytes and macrophages. Keywords: Chemokine; CK9; trout; B lymphocytes; macrophages. x Corresponding author. Tel.þ34 916202300; Fax þ34 916202247. E-mail address: [email protected] (C. Tafalla).
O-072. Characterization of antigen sampling cells in rainbow trout gill epithelium Goshi Kato 1, §, Haruya Miyazawa 1, Takuya Yamaguchi 2, Hidehiro Kondo 1, Motohiko Sano 1, Uwe Fischer 2 1
Tokyo University of Marine Science and Technology, Konan 4-5-7, Minato-ku, Tokyo, 108-8477, Japan 2 Friedrich-Loeffler-Institut, Südufer 10, Greifswald, Insel Riems, 17493, Germany Abstract In mammalian mucosal surfaces, microfold cells (M cells) take up antigens and deliver it to antigen presenting cells in lymphoid tissues such as Payer’s patches. In rainbow trout, a part of gill epithelial cells take up Aeromonas salmonicida subsp. salmonicida (A.s.s.) bacterin during bathvaccination. Antigen-sampling cells in gill epithelium could be stained
Abstracts / Fish & Shellfish Immunology 53 (2016) 58e93
with lectin Ulex europaeus agglutinin 1 (UEA-1) but not with wheat germ agglutinin (WGA); this staining pattern is characteristic for mammalian M cells. In this study, we sorted A.s.s.þ UEA-1þ cells from rainbow trout gill epithelium and performed a comprehensive gene expression analysis using next generation sequencing to characterize the corresponding antigen sampling cells. Rainbow trout (approx. 150 g) were bath-vaccinated with Syto61-stained A.s.s. bacterin. Epithelial cells were isolated from the gills and stained with FITC-labeled UEA-1 and subjected to flow cytometry followed by sorting. Two cell populations, A.s.s.þ UEA-1þ and A.s.s.þ UEA-1- were isolated and total RNA was extracted from each cell population. The cDNA library of each cell population was sequenced using a MiSeq system and gene expression analysis based on the read counts was performed using Trinity assemble software. Flow cytometry revealed that there were two distinct cell populations containing the A.s.s. bacterin in the epithelium: A.s.s.þ UEA-1þ and A.s.s.þ UEA-1-. The percentage of each cell group was 22% in A.s.s.þ UEA-1þ, and 5% in A.s.s.þ UEA-1- population. These two populations also showed different scatter characteristics in flow cytometry, where the A.s.s.þ UEA1þ population exhibited a relatively high side scatter as a measure for pronounced granulation. Remarkably, the A.s.s.þ UEA-1þ population highly expressed annexin which is typical for mammalian M cells, and interleukin (IL)-20 which is typical for mammalian epithelial cells. In contrast, A.s.s.þ UEA-1- cells showed high expression levels of CD83 and IL-12 that are typical for mammalian macrophages and dendritic cells. MHC class II related genes were highly expressed in both of the cell populations from rainbow trout gills. These results suggest that almost of the bacterin are taken up by the UEA-1þ antigen-sampling cells which show properties of epithelial cells during bath-vaccination. In addition, macrophage/dendritic cell lineage may also take part in the uptake of the bacterin.
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O-074. Neutrophil granulocytes in atlantic salmon (Salmo salar L.) are MHCII positive antigen-presenting cells Anita Rønneseth§, Gyri T. Haugland, Lene Gundersen, Harald S. Lunde, Kaja Nordland, Heidrun I. Wergeland Department of Biology, University of Bergen, Norway Abstract In fish, like in mammals, professional antigen presenting cells like dendritic cells, macrophages and B cells are phagocytic cells that ingest, process and present antigens for T cells via MHCII. Neutrophils are professional phagocytic cells with an oxygen-dependent degradation mechanism. By use of a monoclonal antibody against neutrophils in Atlantic salmon, we have shown that the neutrophil granulocytes are MHCII-positive cells. The neutrophils were isolated by magnetic cell sorting and morphological characterization and phenotypic analyses were performed. After in vitro challenge with bacteria, IL12p40, which is essential for proliferation of naïve T cells, were upregulated. Further, Atlantic salmon was bath challenge with the same bacteria as the in vitro challenge experiment and the fraction of neutrophil granulocytes, B cells and MHCII positive cells in primary and secondary immune organs were measured by flow cytometry. Antigen-presentation in fish will be discussed. Keywords: Neutrophils, antigen-presenting cells, bacteria challenge, MHCII x Corresponding author. Tel.: þ47 55584421. E-mail address: [email protected] (A. Rønneseth).
Keywords: antigen up take, antigen sampling cell, mucosal immunity, rainbow trout, next generation sequencer x Corresponding author. Tel: þ81 3 5463 0462. E-mail address: [email protected] (G. Kato).
O-075. Atlantic salmon physiological and immune response to amoebic gill disease and insight into the biology of the amoeba Ottavia Benedicenti 1, 2, §, Chris Secombes 1, Una McCarthy 2, Catherine Collins 2, §
O-073. Characterization of dendritic-like cells generated from hematopoietic spleen cultures from atlantic salmon (Salmo salar L.) Gyri T. Haugland§, Harald S. Lunde, Charlotte F. Asserson, Heidrun I. Wergeland Department of Biology, University of Bergen, Norway Abstract Dendritic cells (DCs) are the main organizers of the immune system in mammals bridging innate and adaptive immunity. DC-like cells are now also identified in teleost fish. At present there is little knowledge of fish DCs properties and functions due to lack of tools to isolate and analyze them. We have developed a protocol for generation of DC-like cells from hematopoietic spleen cultures from Atlantic salmon (Salmo salar L.). In the culture flasks, a stromal layer appeared and after 2 weeks incubation hematopoietic foci were formed. We will describe phenotype and morphology of both adherent and non-adherent MHCIIþ DC-like cells. Upon challenge with bacteria IL12p40, CD83 and CD80/86 were significantly upregulated in the non-adherent DC-like cells. These results support that the cells produced from the hematopoietic cultures are professional antigen cells with potential to activate naïve T-cells belonging to the dendritic cell linage. x Corresponding author. Tel.: þ47 55584445. E-mail address: [email protected] (G.T. Haugland).
1 Scottish Fish Immunology Research Centre, Institute of Biological and Environmental Sciences, University of Aberdeen, Tillydrone Avenue, Aberdeen AB24 2TZ, UK 2 Marine Scotland Science Marine Laboratory, 375 Victoria Rd, Aberdeen AB11 9DB, UK
Abstract Amoebic gill disease (AGD) is an emerging disease in North European Atlantic salmon (Salmo salar) aquaculture caused by the amoeba Paramoeba perurans. Non-optimal environmental conditions such as increasing water temperature may affect AGD progression. To understand the role of predisposing environmental conditions on the biology of the parasite and on the host physiological and immune response, different clonal cultures of P. perurans and AGD infected Atlantic salmon were exposed, respectively, in vitro and in vivo to two different temperatures, 10 C and 15 C. Outputs from the in vitro experiment are 1) an improved understanding of the impact of temperature on amoebae growth rate during conditions that potentially influence disease development and 2) characterisation of changes in bacterial communities at different temperatures for different clonal cultures. Negative binomial analysis in R (R software, version 3.0.1) showed significant differences in growth rate among clonal cultures over time. Moreover, for each clonal culture, the fraction of amoebae in suspension in seawater showed a higher increase over time at 10 C, whereas the fraction of amoebae attached to the malt yeast agar (MYA) showed a higher increase over time at 15 C. 16S MiSeq analysis was also performed to characterise the changes in bacterial communities present in the cultures at the two different temperatures of the in vitro experiment. The host physiological and immune response to AGD infection during predisposing seasonal weather conditions (summer period) is still not well understood. Atlantic salmon smolts were exposed in vivo to the same
Abstracts / Fish & Shellfish Immunology 53 (2016) 58e93 1 Center of Evolutionary and Theoretical Immunology Department of Biology University of New Mexico, New Mexico, USA 2 Research Division Clear Springs Foods Inc, Buhl, Idaho, USA
Abstract Mucosal surfaces are multifunctional barriers that need to orchestrate a variety of physiological functions. One of the mechanisms by which mucosal surfaces carry immunological functions without compromising other physiological roles is the establishment of regional immunity. Regional immunity can be generated by selective expression of chemokines, chemokine receptors and adhesion molecules within mucosal microenvironments. In the olfactory organ of animals, immune responses may result in damage to the olfactory sensory neurons and, as a consequence, compromise olfaction. The olfactory organ of teleost fish is histologically divided into two distinct areas, a sensory or neuroepithelial region in the lateral regions and valleys if the lamellae and a non-sensory region located at the tip of each lamella characterized by presence of goblet cells. Thus, we hypothesized the presence of two unique immune microenvironments corresponding with these two regions in rainbow trout. In line with our hypothesis, we found clusters of CD8þ T cells at the mucosal regions of lamellae whereas few numbers were found in the neuroepithelium. Nasal CD8þ T cells displayed a mucosal phenotype similar to those found in the gut. Laser capture microdissected tip versus lateral samples of control trout olfactory tissue were collected and RTqPCR revealed a higher expression of the chemokine and chemokine receptor CCL19 and CCR7, the adhesion molecules L-selectin, ICAM, VCAM, and integrin b2 and several T cell markers in the mucosal tip compared to the neuroepithelium. Moreover, MHC-II immunostaining indicated that the mucosal tip preferentially participates in antigen presentation within the trout nasal mucosa. Finally, in response to nasal vaccination, cell proliferation occurs in the tip mucosal region and not the neuroepithelium. Our results support the idea of compartmentalized immune responses within the nasal mucosa of teleosts that likely evolved as a sparing mechanism to protect olfactory sensory function. Keywords: Regional immunity, NALT, trout, nasal immunity, mucosal immunity * Corresponding author. Tel.: þ1 505 277 0039. E-mail address: [email protected] (I. Salinas).
O-070. Characterisation and bioactivities of type I interferon subtype E and F in salmonids Thitiya Summathed 1, *, Jun Zou 1, Simon Wadsworth 2, Chris Secombes 1 1
Scottish Fish Immunology Research Centre, Institute of Biological and Environmental Sciences, University of Aberdeen, Tillydrone Avenue, Aberdeen, AB24 2TZ, UK 2 EWOS Innovation A.S., Dirdal, N-4335, Norway Abstract Salmonids are important commercial species in the European aquaculture industry. Many viral diseases can impact on the industry, leading to economic losses. Interferons (IFNs) are cytokines involved in antiviral defence, and study of their regulation and function may shed light on how fish can defend themselves against viral infection. Fish type I IFNs are classified into two groups based on cysteine residues in the mature peptide, namely group I and II IFNs, each of which is further divided into 3 subtypes in salmonids. The latest new subtypes, IFN-e and IFN-f, have only recently been identified and their biological activities have still to be characterised. In this presentation, IFN-e and ef genes in Atlantic salmon (Salmo salar) and their bioactivities were reviewed and compared to previous studies of other type I IFN subtypes (a-d). The type I IFN subtypes showed different expression patterns between the 2 groups in cell lines and primary head kidney cells stimulated with poly I:C and in heart and head kidney of cardiomyopathy syndrome (CMS) infected salmon. Transcript levels could be correlated to the antiviral response and the IFN-e and IFNef
recombinant proteins were shown to induce Mx gene expression which is known to mediate antiviral activity. This data together with the reactivity of specific antibodies to the subtypes provides a better understanding of the antiviral effects of type I IFNs in salmonids. Keywords: Salmonids, type I interferons, bioactivities * Corresponding author. E-mail address: [email protected] (T. Summathed).
O-071. CK9, an ancient rainbow trout chemokine that attracts and regulates B lymphocytes and macrophages Carolina Aquilino 1, Aitor G. Granja 1, Rosario Castro 1, Tiehui Wang 2, Beatriz Abos 1, David Parra 3, Christopher J. Secombes 2, Carolina Tafalla 1, § n en Sanidad Animal (CISA-INIA). Valdeolmos, Centro de Investigacio Madrid, Spain 2 Scottish Fish Immunology Research Centre, School of Biological Sciences, University of Aberdeen, Aberdeen AB24 2TZ, UK 3 Animal Physiology Unit, Department of Cell Biology, Physiology and Immunology, School of Biosciences, Universitat Autonoma de Barcelona, Cerdanyola del Valles, Spain 1
Abstract CK9 is a rainbow trout (Oncorhynchus mykiss) CC chemokine phylogenetically related to mammalian CCL25. Although CK9 is known to be transcriptionally regulated in response to inflammation particularly in mucosal tissues, its functionality has never been revealed. In the current work, we have demonstrated that CK9 has a strong chemotactic capacity for both B cells (IgMþ and IgTþ) and macrophages. Along with its chemotactic capacities, CK9 modulated the MHC II turnover of B lymphocytes and upregulated the phagocytic capacity of both IgMþ cells and macrophages. Although CK9 had no lymphoproliferative effects, it increased the survival of IgTþ lymphocytes. Furthermore, we have established that the chemoattractant capacity of CK9 is strongly increased after pre-incubation of leukocytes with a T-independent antigen, whereas B cell receptor (BCR) cross-linking strongly abrogated their capacity to migrate to CK9, indicating that CK9 preferentially attracts B cells at the steady state or under BCR-independent stimulation. These results point to CK9 being a key regulator of B lymphocyte mucosal trafficking in teleost fish, able to modulate innate functions of teleost B lymphocytes and macrophages. Keywords: Chemokine; CK9; trout; B lymphocytes; macrophages. x Corresponding author. Tel.þ34 916202300; Fax þ34 916202247. E-mail address: [email protected] (C. Tafalla).
O-072. Characterization of antigen sampling cells in rainbow trout gill epithelium Goshi Kato 1, §, Haruya Miyazawa 1, Takuya Yamaguchi 2, Hidehiro Kondo 1, Motohiko Sano 1, Uwe Fischer 2 1
Tokyo University of Marine Science and Technology, Konan 4-5-7, Minato-ku, Tokyo, 108-8477, Japan 2 Friedrich-Loeffler-Institut, Südufer 10, Greifswald, Insel Riems, 17493, Germany Abstract In mammalian mucosal surfaces, microfold cells (M cells) take up antigens and deliver it to antigen presenting cells in lymphoid tissues such as Payer’s patches. In rainbow trout, a part of gill epithelial cells take up Aeromonas salmonicida subsp. salmonicida (A.s.s.) bacterin during bathvaccination. Antigen-sampling cells in gill epithelium could be stained
Abstracts / Fish & Shellfish Immunology 53 (2016) 58e93
with lectin Ulex europaeus agglutinin 1 (UEA-1) but not with wheat germ agglutinin (WGA); this staining pattern is characteristic for mammalian M cells. In this study, we sorted A.s.s.þ UEA-1þ cells from rainbow trout gill epithelium and performed a comprehensive gene expression analysis using next generation sequencing to characterize the corresponding antigen sampling cells. Rainbow trout (approx. 150 g) were bath-vaccinated with Syto61-stained A.s.s. bacterin. Epithelial cells were isolated from the gills and stained with FITC-labeled UEA-1 and subjected to flow cytometry followed by sorting. Two cell populations, A.s.s.þ UEA-1þ and A.s.s.þ UEA-1- were isolated and total RNA was extracted from each cell population. The cDNA library of each cell population was sequenced using a MiSeq system and gene expression analysis based on the read counts was performed using Trinity assemble software. Flow cytometry revealed that there were two distinct cell populations containing the A.s.s. bacterin in the epithelium: A.s.s.þ UEA-1þ and A.s.s.þ UEA-1-. The percentage of each cell group was 22% in A.s.s.þ UEA-1þ, and 5% in A.s.s.þ UEA-1- population. These two populations also showed different scatter characteristics in flow cytometry, where the A.s.s.þ UEA1þ population exhibited a relatively high side scatter as a measure for pronounced granulation. Remarkably, the A.s.s.þ UEA-1þ population highly expressed annexin which is typical for mammalian M cells, and interleukin (IL)-20 which is typical for mammalian epithelial cells. In contrast, A.s.s.þ UEA-1- cells showed high expression levels of CD83 and IL-12 that are typical for mammalian macrophages and dendritic cells. MHC class II related genes were highly expressed in both of the cell populations from rainbow trout gills. These results suggest that almost of the bacterin are taken up by the UEA-1þ antigen-sampling cells which show properties of epithelial cells during bath-vaccination. In addition, macrophage/dendritic cell lineage may also take part in the uptake of the bacterin.
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O-074. Neutrophil granulocytes in atlantic salmon (Salmo salar L.) are MHCII positive antigen-presenting cells Anita Rønneseth§, Gyri T. Haugland, Lene Gundersen, Harald S. Lunde, Kaja Nordland, Heidrun I. Wergeland Department of Biology, University of Bergen, Norway Abstract In fish, like in mammals, professional antigen presenting cells like dendritic cells, macrophages and B cells are phagocytic cells that ingest, process and present antigens for T cells via MHCII. Neutrophils are professional phagocytic cells with an oxygen-dependent degradation mechanism. By use of a monoclonal antibody against neutrophils in Atlantic salmon, we have shown that the neutrophil granulocytes are MHCII-positive cells. The neutrophils were isolated by magnetic cell sorting and morphological characterization and phenotypic analyses were performed. After in vitro challenge with bacteria, IL12p40, which is essential for proliferation of naïve T cells, were upregulated. Further, Atlantic salmon was bath challenge with the same bacteria as the in vitro challenge experiment and the fraction of neutrophil granulocytes, B cells and MHCII positive cells in primary and secondary immune organs were measured by flow cytometry. Antigen-presentation in fish will be discussed. Keywords: Neutrophils, antigen-presenting cells, bacteria challenge, MHCII x Corresponding author. Tel.: þ47 55584421. E-mail address: [email protected] (A. Rønneseth).
Keywords: antigen up take, antigen sampling cell, mucosal immunity, rainbow trout, next generation sequencer x Corresponding author. Tel: þ81 3 5463 0462. E-mail address: [email protected] (G. Kato).
O-075. Atlantic salmon physiological and immune response to amoebic gill disease and insight into the biology of the amoeba Ottavia Benedicenti 1, 2, §, Chris Secombes 1, Una McCarthy 2, Catherine Collins 2, §
O-073. Characterization of dendritic-like cells generated from hematopoietic spleen cultures from atlantic salmon (Salmo salar L.) Gyri T. Haugland§, Harald S. Lunde, Charlotte F. Asserson, Heidrun I. Wergeland Department of Biology, University of Bergen, Norway Abstract Dendritic cells (DCs) are the main organizers of the immune system in mammals bridging innate and adaptive immunity. DC-like cells are now also identified in teleost fish. At present there is little knowledge of fish DCs properties and functions due to lack of tools to isolate and analyze them. We have developed a protocol for generation of DC-like cells from hematopoietic spleen cultures from Atlantic salmon (Salmo salar L.). In the culture flasks, a stromal layer appeared and after 2 weeks incubation hematopoietic foci were formed. We will describe phenotype and morphology of both adherent and non-adherent MHCIIþ DC-like cells. Upon challenge with bacteria IL12p40, CD83 and CD80/86 were significantly upregulated in the non-adherent DC-like cells. These results support that the cells produced from the hematopoietic cultures are professional antigen cells with potential to activate naïve T-cells belonging to the dendritic cell linage. x Corresponding author. Tel.: þ47 55584445. E-mail address: [email protected] (G.T. Haugland).
1 Scottish Fish Immunology Research Centre, Institute of Biological and Environmental Sciences, University of Aberdeen, Tillydrone Avenue, Aberdeen AB24 2TZ, UK 2 Marine Scotland Science Marine Laboratory, 375 Victoria Rd, Aberdeen AB11 9DB, UK
Abstract Amoebic gill disease (AGD) is an emerging disease in North European Atlantic salmon (Salmo salar) aquaculture caused by the amoeba Paramoeba perurans. Non-optimal environmental conditions such as increasing water temperature may affect AGD progression. To understand the role of predisposing environmental conditions on the biology of the parasite and on the host physiological and immune response, different clonal cultures of P. perurans and AGD infected Atlantic salmon were exposed, respectively, in vitro and in vivo to two different temperatures, 10 C and 15 C. Outputs from the in vitro experiment are 1) an improved understanding of the impact of temperature on amoebae growth rate during conditions that potentially influence disease development and 2) characterisation of changes in bacterial communities at different temperatures for different clonal cultures. Negative binomial analysis in R (R software, version 3.0.1) showed significant differences in growth rate among clonal cultures over time. Moreover, for each clonal culture, the fraction of amoebae in suspension in seawater showed a higher increase over time at 10 C, whereas the fraction of amoebae attached to the malt yeast agar (MYA) showed a higher increase over time at 15 C. 16S MiSeq analysis was also performed to characterise the changes in bacterial communities present in the cultures at the two different temperatures of the in vitro experiment. The host physiological and immune response to AGD infection during predisposing seasonal weather conditions (summer period) is still not well understood. Atlantic salmon smolts were exposed in vivo to the same