repertoires for IgM, IgD and IgT, respectively, with a few ampliﬁed m and s junctions, suggesting the presence of IgM- and IgT-secreting cells in the spleen. In infected animals, we detected complex and highly diverse IgM responses involving all VH subgroups, and dominated by a few large public and private clones. A lower number of robust clonal responses involving only a few VH were detected for the mucosal IgT, indicating that both IgM+ and IgT+ spleen B cells responded to systemic infection but at different degrees. In contrast, the IgD response to the infection was faint. Although ﬁsh IgD and IgT present different structural features and evolutionary origin compared to mammalian IgD and IgA, respectively, their implication in the B-cell response evokes these mouse and human counterparts. Thus, it appears that the general properties of antibody responses were already in place in common ancestors of ﬁsh and mam- mals, and were globally conserved during evolution with possible func- tional convergences. * Corresponding author. E-mail address: pierre.boudinot@jouy.inra.fr (P. Boudinot) O-203. Immunomodulatory effects of Ganoderma lucidum and virgin coco- nut oil in Nile tilapia artiﬁcially-infected with Aeromonas hydrophila E.S. Catap 1, *, x , M.R. Jimenez 1, 2, x , S.C.O. Liquido 1, x , M.A.K.B. Cabujat 1, x , M.M. Sadaya 1, x . 1 Institute of Biology, College of Science, University of the Philippines, Diliman, Quezon 1101, Philippines; 2 Natural Sciences Research Institute, College of Science, University of the Philippines, Diliman, Quezon 1101, Philippines Abstract Oreochromis niloticus or Nile tilapia, is the second most important aqua- culture species in the Philippines. High demands of ﬁsh products and intensiﬁcation of pond cultures have subjected these ﬁshes to various environmental stress that lead to infectious pathogens, such as Aero- monas hydrophila. Hence, immunomodulators such as Ganoderma lucidum and virgin coconut oil (VCO) have been considered as means to control infection and prevent ﬁsh mortality. The use of immunostimulators ne- gates the use of expensive synthetic therapeutants and antibiotics to ﬁght infectious diseases. Two separate experiments were undertaken to assess the effects of powdered G. lucidum and VCO in O. niloticus injected with A. hydrophila. For each experiment, three tanks of Nile tilapia were prepared which corresponds with the experimental groups. The ﬁrst group of ﬁsh served as the negative control and were injected with phosphate-buff- ered saline; the second group served as the positive control ﬁsh which injected with 0.1 ml A. hydrophila (1 x 10 6 cfu/ml); and the third group of ﬁsh served as the treatment set-up wherein the ﬁsh were either fed with pellets coated with 0.12% G. lucidum or with VCO-coated pellets, and then injected with A. hydrophila. Fish were infected with A. hydrophila after 30 days of feeding with either VCO or G. lucidum. Blood samples, head kid- neys phagocytes and splenic lymphocytes were harvested at days 8 and 15 post-infection and were used to assay phagocytosis activity, lysozyme levels, and reactive oxygen species (ROS) production and lymphocyte proliferation were measured at day 15 p.i. Both G. lucidum- and VCO- supplemented groups of ﬁsh signiﬁcantly induced higher phagocytic ac- tivity than the positive control ﬁsh at days 8 and 15 p.i. The lysozyme levels showed the same pattern, but the difference between the two groups was not statistically signiﬁcant both at days 8 and days 15 p.i. Signiﬁcantly higher ROS was produced by head kidney macrophages from G. lucidum treated ﬁsh compared with the positive control ﬁsh. Higher ROS production was also obtained from VCO treated ﬁsh but this was not signiﬁcantly different from the positive control data. At day 15 p.i., lower T and B lymphocyte proliferation was obtained in G. lucidum and VCO treated O. niloticus compared with their respective positive control ﬁsh. The results showed that oral feeding of either powdered G. lucidum or VCO enhanced some nonspeciﬁc immune responses, but both treatments did not induce proliferation of both T and B lymphocytes. Further studies are needed to validate the immunomodulatory roles of these natural bioactive products. * Corresponding author. E-mail address: elanacatap@yahoo.com (E.S. Catap) xThese authors have contributed equally to this work. O-452. A tour through high pressure and atmospheric ecosystems: Immune responses in hydrothermal vent Bathymodiouls azoricus and Mediter- ranean Mytilus galloprovincialis mussels T. Cerqueira * , I. Martins, R. Bettencourt. IMAR Centre, Department of Oceanography and Fisheries, University of the Azores, Portugal Abstract Deep-sea hydrothermal vents are considered hostile environments by normal life standards. The toxic chemical nature of vent surrounding could seemingly be deleterious to the animals, yet among ﬂuids rich in sulfur compounds, methane and high concentration of trace metals, animals have developed successful strategies to cope with adverse physical and chemical environmental conditions. On the other hand, coastal mussels have to embrace hydrodynamic stress which is the main disturbance factor inﬂuencing intertidal communities. Tidal var- iations condition mussel's food and oxygen supply. Besides the inherent stressful characteristics on both environments, hydrostatic pressure is one of the most important parameter that distinguishes the deep-sea from coastal ecosystems. The extraordinary physiological adaptation to high pressure conditions raises interesting biological questions. In the present study we characterize the expression and regulation of immune genes in both B. azoricus and M. galloprovincialis individuals under different experimental pressurized conditions. Hy- drothermal vent B. azoricus individuals were acclimatized for 3 months to atmospheric pressure in LabHorta aquarium system and subjected to high hydrostatic pressure (80 bar) using the IPOCAMP system for 2 days. The Mediterranean M. galloprovincialis individuals were also subjected to 80 bar of hydrostatic pressure for the same period of days. Patterns of gene expression, in the gills of both species, were assessed by RT-qPCR. In parallel, protein proﬁles and proteolytic activity expressed in experimental individuals were evaluated using SDS-PAGE and zymography techniques. This study unveils the immune genes modulated to endure high hydrostatic pressure as well as the molec- ular mechanisms involved in speciﬁc adaptation processes to two distinct natural habitats. * Corresponding author. E-mail address: mcerqueira@uac.pt (T. Cerqueira) O-416. Effect of seeds of Achyranthes aspera on the immune responses and expression of some immune-related genes in Catla catla R. Chakrabarti 1, *, x , P.K. Srivastava 1, x , N. Verma 1 , J.G. Sharma 2, x . 1 Aqua Research Lab, Department of Zoology, University of Delhi, Delhi 110007, India; 2 Department of Biotechnology, Delhi Technological University, Delhi 110042, India Abstract Evaluation of functional mechanism of immunostimulant in ﬁsh is essential for realizing their therapeutic potential. The impact of dietary supplementation of seeds of Achyranthes aspera on the immune system of Catla catla was studied. Catla (37.736.7 g) were fed with test diets containing 0.5 (D1) and 1.0% (D2) seeds of Achyranthes aspera and control diet (D3) for 50 days in three replicated tanks. Then ﬁsh were immunized with heat-killed Aeromonas hydrophila. Supplementation of Abstracts / Fish & Shellﬁsh Immunology 34 (2013) 1635–1691 1644