Opisthorchis viverrini is the only liver fluke that has been proved to be associated with cholangiocarcinoma (CCA). However, the mechanisms by which O. viverrini participates in the carcinogenesis of CCA are still unclear. To understand the biology and host-parasite interaction related to O. viverrini infection, gene expression profiling of this parasite is required. Here, we constructed the first 5' serial analysis of gene expression (5' SAGE) library of the adult O. viverrini and matched the data with the public data of O. viverrini, Clonorchis sinensis and other related Platyhelminthes and Nematodes. We obtained 12,401 unique tag sequences, of which 6515 (53%) could be matched with the 3419 transcript sequences. The two most abundant tag sequences were vitelline B precursor protein and myoglobin. Often several transcription start sites (TSS) were observed for one transcript. This finding may reflect the dynamic nature of transcriptional initiation events of O. viverrini genes in vivo.