1654 Abstracts / Fish & Shellfish Immunology 34 (2013) 1635–1691 Abstract Differential complementarity of immune receptors and pathogene surface markers is a key recognition mechanism in all immune systems. The cell surface receptor Down syndrome cell adhesion molecule (Dscam) subjected to mutually exclusive splicing in Arthropoda shows an extraordinary degree of potential immune recognition specificity. We have tested the hypothesis on the existence of similar polymorphism in the homologous receptors from sister-clade protostomian animals, Mollusca. Using degenerative RT-PCR, two new members of Dscam family from bivalve mollusks, Mytilus edulis (MeDscam) and Crassostrea gigas (CgDscam) were cloned from nervous ganglia mRNA and their differential expression was estimated in immune cells. The expression of MeDscam and CgDscam in haemocytes is weak, constitutive and is not modulated by immune inducers neither in vivo nor in vitro experiments. It is important to note that the same approach used for discovering Dscam in gastropod Littorina littorea has proven unsuccessful. We did not found an expression of this receptor in the common periwinkle haemocytes. The following deep sequencing of the L. littorea immune cells transcriptome (NGS; in preparation) evidenced the lack of Dscam transcripts in littorina immune cells. Based on our sequencing results, we did not observe diversity in the Dscam transcripts in ganglia and haemocytes of M. edulis and C. gigas. Moreover, the screening of oyster genome and genomes of two gastropod mollusks, Lottia gigantea and Aplysia californica, revealed no their Dscam homologues genes do contain the hypervariable exon arrays and all exons are found at unique positions. This type of gene structure leaves no chance for the generation of multiple splice variants. Thus, we can conclude that the immune function of Dscam pattern recognition receptor based on the vast repertoire of isoforms is an evolutionary innovation developed exclusively in the Arthropoda lineage and could not be attributed to representatives of Mollusca. Extrapolating our data, we predict that, even within the phylum Mollusca the functions of Dscam receptor in immune system evolved differently. In Gastropoda, Dscam is not involved in the immune mechanisms, while in Bivalvia, retaining the ancient original function, acts as cell-to-cell adhesion molecule, such as is involved in hemocytes aggregation. * Corresponding author. E-mail address: agorbushin@gmail.com (A.M. Gorbushin) O-428. Immune gene expression profiling of proliferative kidney disease in rainbow trout Oncorhynchus mykiss reveals a dominance of anti-inflammatory, antibody and Th cell-like activities B. Gorgoglione, T. Wang, C.J. Secombes, J.W. Holland*. Scottish Fish Immunology Research Centre, School of Biological Sciences, University of Aberdeen, Tillydrone Avenue, Aberdeen AB24 2TZ, UK Abstract The myxozoan Tetracapsuloides bryosalmonae is the causative agent of proliferative kidney disease (PKD) targeting primarily the kidney of infected fish where it causes a chronic lymphoid immunopathology. Although known to be associated with suppression of some cellular aspects of innate immunity and a prominent lymphocytic hyperplasia, there remains a considerable knowledge gap in our understanding of the underlying immune mechanisms driving PKD pathogenesis. To provide further insights, the expression profiles of a panel of innate / inflammatory and adaptive immune molecules were examined in rainbow trout Oncorhynchus mykiss following a natural exposure to the parasite. Relative to controls, fish with early to advanced stages of kidney pathology exhibited up-regulation of the inflammatory cytokines IL-6 and IL-11, although remaining refractory towards genes indicative of macrophage activity. Antimicrobial peptides (AMPs) and anti-inflammatory markers, including cathelicidin and IL-10 were markedly up-regulated during clinical disease. Up-regulation of adaptive immune molecules, including cell markers and antibody genes reflect the lymphocytic dominance of this disease and the likely importance of lymphocyte subsets in PKD pathogenesis. Up-regulation of TH-type response genes and transcription factors implies that T. bryosalmonae may elicit a complex interplay between TH subsets. This work, for the first time in the study of fish-myxozoan interactions, suggests that PKD pathogenesis is shaped by an anti-inflammatory phenotype, a profound B cell / antibody response and dysregulated TH cell-like activities. A better understanding of the functional roles of fish immune cells and molecules in PKD pathogenesis may facilitate future development of control measures against this disease. * Corresponding author. E-mail address: j.holland@abdn.ac.uk (J.W. Holland) O-196. Isolation and identification of lymphoid subpopulations from Salmo salar N. Halçartégaray 1, 2, *, J. Neira 1, P. Reyes 1, V. Wilhelm 1, 2, M.R. Bono 2, M. Rosemblatt 1, 2, 3. 1 Fundación Ciencia & Vida, Santiago, Chile; Facultad de Ciencias Biológicas, Universidad Andrés Bello, Santiago, Chile; 3 Facultad de Ciencias Universidad de Chile, Santiago, Chile 2 Abstract Salmon lymphoid cells are poorly characterized, mainly due to the lack of monoclonal antibodies (mAbs) able to identify them. Here we report on the isolation and characterization of different subpopulations of lymphoid cells by the generation and use of specific monoclonal antibodies. For the preparation of monoclonal antibodies, mice were immunized with head kidney mononuclear cells. Hybridomas that recognized live head kidney cells were selected by FACS. Lymphoid cells from Salmo salar were further analyzed with our mAbs and a mAb to salmonid IgM. We obtained five hybridomas that recognize different subpopulations of lymphoid cells from salmon head kidney and spleen. Use of the hybridomas NH2.10, NH2.16 and NH2.18 in conjuction with the a-IgM monoclonal antibody allowed for the identification of three different cell populations: double-positive cells, double-negative cells, and NHpos/ IgMneg cells. These subpopulations were isolated by cell sorting with a better than 96% purity and analyzed by a Real-Time quantitative PCR assay. The qPCR analyses of the isolated subpopulations demonstrated a differential expression patterns of relevant genes (CD4, CD3, CD8, TCR, IL-2, IL10, IgM, IFNg, MHC-II), allowing for the recognition of T cells among the double-negative subpopulations, and of putative MHCpos antigen presenting cells (APC) among the NH2.10pos/IgMneg subpopulation. These results show that the mAbs prepared against live salmon kidney mononuclear cells can be used to identify and separate subpopulations of lymphoid T cells and APCs. We aim at using isolated cells in activation assays in vitro. * Corresponding author. E-mail address: nicole.halcartegaray@gmail.com (N. Halçartégaray) O-477. Efficiency of different immunostimulants on immune response of striped catfish (Pangasianodon hypophthalmus) B.T. Hang 1, 2, N.T. Phuong 2, P. Kestemont 1, *. 1 Research Unit in Environmental and Evolutionary Biology, University of Namur, rue de Bruxelles 61, B-5000 Namur, Belgium; 2 College of Aquaculture and Fisheries, Can Tho University, Campus 2, Can Tho, Viet Nam Abstract Different immunostimulants were investigated to detect changes in the activity of non-specific immune response and disease resistance of striped catfish (Pangasianodon hypophthalmus). In current experiment, striped catfish were injected with five immunostimulants such as beta-glucan (10mg/kg fish), bovine lactoferrin (500mg/kg fish), chitosan (6mg/kg fish), levamisole (5mg/kg fish) and vitamin C (1,000 mg/kg fish). The plasma cortisol and glucose were measured as stress biomarkers, the respiratory