title: Identification and analysis of differentially expressed genes in immune tissues of Atlantic cod stimulated with formalin-killed, atypical Aeromonas salmonicida.
abstract: Physiological changes, elicited in animal immune tissues by exposure to pathogens, may be studied using functional genomics approaches. We created and characterized reciprocal suppression subtractive hybridization (SSH) cDNA libraries to identify differentially expressed genes in spleen and head kidney tissues of Atlantic cod (Gadus morhua) challenged with intraperitoneal injections of formalin-killed, atypical Aeromonas salmonicida. Of 4,154 ESTs from four cDNA libraries, 10 genes with immune-relevant functional annotations were selected for QPCR studies using individual fish templates to assess biological variability. Genes confirmed by QPCR as upregulated by A. salmonicida included interleukin-1 beta, interleukin-8, a small inducible cytokine, interferon regulatory factor 1 (IRF1), ferritin heavy subunit, cathelicidin, and hepcidin. This study is the first large-scale discovery of bacteria-responsive genes in cod and the first to demonstrate upregulation of IRF1 in fish immune tissues as a result of bacterial antigen stimulation. Given the importance of IRF1 in vertebrate immune responses to viral and bacterial pathogens, the full-length cDNA sequence of Atlantic cod IRF1 was obtained and compared with putative orthologous sequences from other organisms. Functional annotations of assembled SSH library ESTs showed that bacterial antigen stimulation caused changes in many biological processes including chemotaxis, regulation of apoptosis, antimicrobial peptide production, and iron homeostasis. Moreover, differences in spleen and head kidney gene expression responses to the bacterial antigens pointed to a potential role for the cod spleen in blood-borne pathogen clearance. Our data show that Atlantic cod immune tissue responses to bacterial antigens are similar to those seen in other fish species and higher vertebrates.
authors.affiliation: Ocean Sciences Centre, Memorial University of Newfoundland, St. John's, Newfoundland.
authors.forename: Charles Y
authors.initials: CY
authors.lastname: Feng
authors.forename: Stewart C
authors.initials: SC
authors.lastname: Johnson
authors.forename: Tiago S
authors.initials: TS
authors.lastname: Hori
authors.forename: Marlies
authors.initials: M
authors.lastname: Rise
authors.forename: Jennifer R
authors.initials: JR
authors.lastname: Hall
authors.forename: A Kurt
authors.initials: AK
authors.lastname: Gamperl
authors.forename: Sophie
authors.initials: S
authors.lastname: Hubert
authors.forename: Jennifer
authors.initials: J
authors.lastname: Kimball
authors.forename: Sharen
authors.initials: S
authors.lastname: Bowman
authors.forename: Matthew L
authors.initials: ML
authors.lastname: Rise
mesh_terms: D048409:Aeromonas salmonicida / Q000276:immunology*; D000595:Amino Acid Sequence; D000818:Animals; D001428:Bacterial Vaccines / Q000008:administration & dosage / Q000276:immunology; D001483:Base Sequence; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D029941:Fish Proteins / Q000235:genetics; D005557:Formaldehyde; D048389:Gadus morhua / Q000145:classification / Q000235:genetics*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects / Q000276:immunology; D015723:Gene Library; D007274:Injections, Intraperitoneal; D050856:Interferon Regulatory Factor-1 / Q000145:classification; D007668:Kidney / Q000378:metabolism*; D008969:Molecular Sequence Data; D010802:Phylogeny; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013154:Spleen / Q000378:metabolism*; D014613:Vaccines, Attenuated / Q000008:administration & dosage / Q000276:immunology
chemical_list: D001428:Bacterial Vaccines; D018076:DNA, Complementary; D029941:Fish Proteins; D050856:Interferon Regulatory Factor-1; D014613:Vaccines, Attenuated; D005557:Formaldehyde