Establishment of a soft shell turtle, Pelodiscus sinensis, embryo primary cell culture for studies of soft shell turtle poxvirus-like virus replication and characteristics

Abstract

The aim of this study was to establish a soft shell turtle primary culture for replication of the isolated poxvirus-like virus. To date, the primary culture cells have been successfully subcultured for more than 25 passages in a T-25 cell culture flask. Based on morphology studies, these cells were identified as mostly fibroblast cells. The results of this study showed that the L15 medium was the optimal culture medium for these cells. Interestingly, the soft shell turtle primary cells grew faster at 37°C than at temperatures below 30°C. The lysis cellular pathogenic effects (CPE) presented on the monolayer cell culture which has been inoculated with 200 μL of the tested isolated poxvirus-like viral suspension (1 × 108 TCID50/ml) on the 2nd postinoculation day. The virulence of poxvirus-like virus was significantly inhibited by 5-iodo-2’-deoxyuridine, but 5-bromo-2’-deoxyuridine had no significant effect. According to the acridine orange staining results, the virus-infected primary cells were yellow-green in color, indicating that the poxvirus-like virus was a double-stranded DNA virus. The set primary culture cells were highly capable of being successfully infected with the soft shell turtle poxvirus-like virus. This study may be the first study to determine the optimal primary culture for studying viral infections in soft shell turtle embryos. Key words: Pox-like virus, soft shell turtle, reptile, primary cell culture, replication.