Establishment of cell culture systems from penaeid shrimp and their susceptibility to white spot disease and yellow head viruses.

Monolayer cultures were established from ovary, heart, lymphoid tissue and peripheral hemocytes of penaeid shrimps including Penaeus monodon, P. japonicus and P. penicillatus. The most favorable conditions for the culture of penaeid shrimp cells in vitro was in CMRL and L-15 tissue culture media when used within an osmolarity range of 620--760 mmol/kg. The optimal maintenance temperature was 25 degrees C for tissues of P. japonicus and 28 degrees C for tissues of P. monodon and P. penicillatus. Among the four tissues tested, lymphoid tissue, or 'Oka organ', was superior to the other tissues for the formation of confluent cell monolayers. Cell cultures from lymphoid tissue and ovary have been subcultured up to three times. When peripheral hemocytes and heart were cultured, a maximum survival of 4 days was obtained. In contrast, cell cultures derived from ovary and lymphoid tissue were maintained alive for at least 20 days in appropriate culture systems. Neither confluent cell sheet nor adherence of cells was obtained in cultivation of hepatopancreas using the present culture systems. The results obtained from the present study also revealed that ovary extract, muscle extract and lobster hemolymph enhanced the survival of the cultured cells of penaeid shrimp in vitro. When the 'Oka organ' cell monolayer was incubated with either white spot disease virus (WSDV) or yellow head virus (YHV), no cytopathic effect (CPE) was obtained. However, at 5--7 days after establishment, significant CPE (a few foci) was observed in cell monolayers derived from WSDV- and YHV-infected Oka tissue. By electron microscopy, virions of WSDV and YHV were observed in the nuclei and cytoplasm of cultured cells. The CPE foci developed further with increased incubation time.