Eudiplozoon nipponicum (Polyopisthocotylea: Diplozoidae): transcriptome and secretome analyses of hematophagous fish parasite

Two sequencing methods (454/Roche and Illumina MiSeq platform) and mass spectrometry analysis (HPLC MS/MS) were used. Total RNA from a few E. nipponicum adults, in the form of cDNA, was sequenced and Illumina and 454/Roche raw reads were processed and assembled separately using specific bioinformatics tools. Based on statistical evaluation, selected assembly variants were polled and filtered, in order to get high quality transcriptomics sequences for further annotation, into final contaminant-free 37,062 transcripts with corresponding translated proteins. Totally 19,539 (52.7 %) transcripts were homologous to some record in used public sequence databases (BLASTn and BLASTp algorithms were used, E-value cut-off: 1e-5) and 18,556 (50.1 %) of them were linked with sequences related to organisms only in phylum Platyhelminthes deposited in UniProtKB/TrEMBL database. All transcripts were deeply annotated by established approach. Excretory-secretory products (ESP) – protein compounds present in parasite secretome, were obtained from ~100 living adults. HPLC MS/MS analysis of peptide mixture were done using RSLCnano System connected to Orbitrap Elite Hybrid spectrometer after tryptic digestion. The analysis of the mass spectrometric raw data was carried out using the Proteome Discoverer software tool with Mascot search engine utilisation and finally, 1,033 transcripts were identified in secretome.