Full length article Comparative study of immune responses in the deep-sea hydrothermal vent mussel Bathymodiolus azoricus and the shallow- water mussel Mytilus galloprovincialis challenged with Vibrio bacteria Eva Martins a, b , Ant  onio Figueras c , Beatriz Novoa c , Ricardo Serr ~ ao Santos a, b , Rebeca Moreira c , Raul Bettencourt b, d, * a Department of Oceanography and Fisheries, University of the Azores (DOP/UAç), Rua Prof. Doutor Frederico Machado, 9901-862 Horta, Portugal b IMAR Institute of Marine Research and LARSyS Laboratory of Robotics and Systems in Engineering and Science, 9901-862 Horta, Azores, Portugal c Instituto de Investigaciones Marinas, IIM e CSIC. Eduardo Cabello, 6, 36208 Vigo, Spain d MARE-Marine and Environmental Science Center, University of the Azores, 9901-862 Horta, Azores, Portugal article info Article history: Received 23 April 2014 Received in revised form 8 July 2014 Accepted 9 July 2014 Available online 1 August 2014 Keywords: Bathymodiolus azoricus Mytilus galloprovincialis Bacterial challenges Gene expression HPLC-ESI-MS/MS abstract The deep-sea hydrothermal vent mussel Bathymodiolus azoricus and the continental European coast Mytilus galloprovincialis are two bivalves species living in highly distinct marine habitats. Mussels are filter-feeding animals that may accumulate rapidly bacteria from the environment. Contact with microorganism is thus inevitable during feeding processes where gill tissues assume a strategic importance at the interface between the external milieu and the internal body cavities promoting in- teractions with potential pathogens during normal filtration and a constant challenge to their immune system. In the present study B. azoricus and M. galloprovincialis were exposed to Vibrio alginolyticus, Vibrio anguillarum and Vibrio splendidus suspensions and to a mixture of these Vibrio suspensions, in order to ascertain the expression level of immune genes in gill samples, from both mussel species. The immune gene expressions were analyzed by means of quantitative-Polymerase Chain Reaction (qPCR). The gene expression results revealed that these bivalve species exhibit significant expression differences between 12 h and 24 h post-challenge times, and between the Vibrio strains used. V. splendidus induced the strongest gene expression level in the two bivalve species whereas the NF-kB and Aggrecan were the most significantly differentially expressed between the two mussel species. When comparing exposure times, both B. azoricus and M. galloprovincialis showed similar percentage of up-regulated genes at 12 h while a marked increased of gene expression was observed at 24 h for the majority of the immune genes in M. galloprovincialis. This contrasts with B. azoricus where the majority of the immune genes were down-regulated at 24 h. The 24 h post-challenge gene expression results clearly bring new evidence supporting time-dependent transcriptional activities resembling acute phase-like responses and different immune responses build-up in these two mussel species when challenged with Vibrio bacteria. High Pressure Liquid Chromatography (HPLC)-Electrospray ionization mass spectrometry (ESI-MS/MS) analyses resulted in different peptide sequences from B. azoricus and M. galloprovincialis gill tissues suggesting that naïve animals present differences, at the protein synthesis level, in their natural envi- ronment. B. azoricus proteins sequences, mostly of endosymbiont origin, were related to metabolic, energy production, protein synthesis processes and nutritional demands whereas in M. galloprovincialis putative protein functions were assumed to be related to structural and cellular integrity and signaling functions. © 2014 Elsevier Ltd. All rights reserved. 1. Introduction The invertebrate immune system distinguishes self from non- self, resulting in physiological responses mediated through cellular and humoral processes that effectively fight against * Corresponding author. MARE-Marine and Environmental Science Center, Uni- versity of the Azores, 9901-862 Horta, Azores, Portugal. Tel.: þ351 292 200 400. E-mail addresses: raul@uac.pt, bettencourt.raul@gmail.com (R. Bettencourt). Contents lists available at ScienceDirect Fish & Shellfish Immunology journal homepage: www.elsevier.com/locate/fsi http://dx.doi.org/10.1016/j.fsi.2014.07.018 1050-4648/© 2014 Elsevier Ltd. All rights reserved. Fish & Shellfish Immunology 40 (2014) 485e499