Submitted to: Biomed Central (BMC) Genomics
Publication Type: Peer reviewed journal
Publication Acceptance Date: 6/18/2007
Publication Date: 6/18/2007
Citation: Abernathy, J.W., Xu, P., Li, P., Xu, D., Kucuktas, H., Klesius, P.H., Arias, C., Liu, Z. 2007. Generation and analysis of expressed sequence tags from the ciliate protozoan parasite Ichthyophthirius multifiliis. Biomed Central (BMC) Genomics. 8:176 pgs. 1-10. Interpretive Summary: Parasite Ichthyophthirius (Ich) infects most species of fresh water fish world wide, causing high mortalities of fish, and leads to heavy economic loss in aquaculture. A genomic resource for large-scale studies of this parrasite has been lacking. To study gene expression involved in Ich virulence, this study generated expressed sequence tags (ESTs) for the analysis of global gene expression in this parasite. We sequenced 10,368 EST clones using a normalized complementary DNA library made from pooled samples of the parasite at three life-cycle stages, and generated 9,769 sequences (94.2% success rate). Post-sequencing processing lead to 8,432 high quality sequences. This set of ESTs represents a significant proportion of the Ich gene information, and provides a material basis for the development of microarrays (DNA chips) useful for gene expression studies. These results are critically important for understanding gene expression on development, pathogenesis, and virulence of the parasite. This study will aid in developing methods to prevent or control this severe fish parasite and greatly reducing the economic loss in aquaculture.
Technical Abstract: The ciliate protozoan Ichthyophthirius multifiliis (Ich) is an important parasite of freshwater fish that causes 'white spot disease' leading to significant losses. A genomic resource for large-scale studies of this parasite has been lacking. To study gene expression involved in Ich pathogenesis and virulence, our goal was to generate expressed sequence tags (ESTs) for the development of a powerful microarray platform for the analysis of global gene expression in this species. Here, we initiated a project to sequence and analyze over 10,000 ESTs. We sequenced 10,368 EST clones using a normalized cDNA library made from pooled samples of the trophont, tomont, and theront life-cycle stages, and generated 9,769 sequences (94.2% success rate). Post-sequencing processing lead to 8,432 high quality sequences. Clustering analysis of these ESTs allowed identification of 4,706 unique sequences containing 976 contigs and 3,730 singletons. These unique sequences represent over two million base pairs (~10% of Plasmodium falciparum genomes, a phylogenetically related protozoan). BLASTX searches produced 2,518 significant (E-value<10-5) hits and further Gene Ontology (GO) analysis annotated 1,008 of these genes. The ESTS were analyzed comparatively against the genomes of the related protozoa Tetrahymena thermophila and P. falciparum, allowing putative identification of additional genes. All the EST sequences were deposited by dbEST in GenBank (GenBank: EG957858-EG966289). Gene discovery and annotations are presented and discussed. This set of ESTs represents a significant proportion of the Ich transcriptome, and provides a material basis for the development of microarrays useful for gene expression studies concerning Ich development, pathogenesis, and virulence.