Transcriptomics of in vitro immune-stimulated hemocytes from the Manila clam Ruditapes philippinarum using high-throughput sequencing

• Published in: PloS one Vol. 7; no. 4; p. e35009
• Main Authors: Moreira, Rebeca, Balseiro, Pablo, Planas, Josep V, Fuste, Berta, Beltran, Sergi, Novoa, Beatriz, Figueras, Antonio
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 19.04.2012; Public Library of Science (PLoS)
• Subjects: Acids; Agriculture; Analysis; Animals; Apoptosis; Apoptosis - genetics; Apoptosis - immunology; Automation; Biology; Bivalvia; Bivalvia - genetics; Bivalvia - immunology; Clams; Cloïsses; Communicable diseases; Communicable diseases in animals; Complement Activation - genetics; Consortia; DNA microarrays; DNA, Bacterial - immunology; Economic impact; Expressed Sequence Tags; Gene expression; Gene sequencing; Genes; Genetic markers; Genomes; Genomics; Hemocytes; Hemocytes - drug effects; Hemocytes - immunology; High-Throughput Nucleotide Sequencing; Immune response; Immune system; Immune System - metabolism; Immunologic Factors - pharmacology; Immunotherapy; Infectious diseases; Lipopolysaccharides - pharmacology; Malalties infeccioses en els animals; Molecular modelling; Molecular Sequence Annotation; Mollusks; Next-generation sequencing; Ontology; Peptidoglycan - immunology; Poly I-C - pharmacology; Receptors, Pattern Recognition - genetics; Receptors, Pattern Recognition - immunology; Resposta immunitària; Ribonucleic acid; RNA; Ruditapes philippinarum; Sequence Analysis, DNA; Signal transduction; Signal Transduction - genetics; Signal Transduction - immunology; Signaling; Taxonomy; Technology application; Transcriptome - immunology; Vibrio - genetics; Vibrio - immunology; Spain
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0035009

The Manila clam (Ruditapes philippinarum) is a worldwide cultured bivalve species with important commercial value. Diseases affecting this species can result in large economic losses. Because knowledge of the molecular mechanisms of the immune response in bivalves, especially clams, is scarce and fragmentary, we sequenced RNA from immune-stimulated R. philippinarum hemocytes by 454-pyrosequencing to identify genes involved in their immune defense against infectious diseases. High-throughput deep sequencing of R. philippinarum using 454 pyrosequencing technology yielded 974,976 high-quality reads with an average read length of 250 bp. The reads were assembled into 51,265 contigs and the 44.7% of the translated nucleotide sequences into protein were annotated successfully. The 35 most frequently found contigs included a large number of immune-related genes, and a more detailed analysis showed the presence of putative members of several immune pathways and processes like the apoptosis, the toll like signaling pathway and the complement cascade. We have found sequences from molecules never described in bivalves before, especially in the complement pathway where almost all the components are present. This study represents the first transcriptome analysis using 454-pyrosequencing conducted on R. philippinarum focused on its immune system. Our results will provide a rich source of data to discover and identify new genes, which will serve as a basis for microarray construction and the study of gene expression as well as for the identification of genetic markers. The discovery of new immune sequences was very productive and resulted in a large variety of contigs that may play a role in the defense mechanisms of Ruditapes philippinarum.