Characterization of white spot syndrome virus replication in in vitro-cultured haematopoietic stem cells of freshwater crayfish, Pacifastacus leniusculus.

Replication of White spot syndrome virus (WSSV) was investigated in haematopoietic cells (hpt cells) derived from haematopoietic tissue (hpt) of freshwater crayfish, Pacifastacus leniusculus. Temperature and type of inoculum for virus replication were studied. The cell culture remained viable at a wide range of temperatures ranging from 4 to 25 degrees C. WSSV replicated in cells, as evidenced by in situ hybridization, RT-PCR and by the presence of virions visualized with an electron microscope. Moreover, the results showed that the infectivity of WSSV to hpt cells is dependent on temperature and a supplemented growth factor (cytokine) astakine. WSSV replicated more rapidly at higher temperatures than at lower temperatures. No virus replication was observed at 4 degrees C. Detectable WSSV-infected cells were present as early as 36 h post-inoculation, demonstrated by in situ hybridization or RT-PCR of VP28 expression at 25 degrees C. Hpt cells can survive a few weeks at 25 or 16 degrees C and longer than several months at 4 degrees C.