Attempts at producing a hybridised Penaeus mondon cell line by cellular fusion

The lack of a standardised system for the isolation, identification and purification of prawn viruses, is a major obstacle to the control of viruses in penaeid aquaculture. To date, spontaneous and induced transformation of somatic penaeid cells has failed. Hybrid cells with the aim of supporting the growth of penaeid viruses were created using polyethylene glycol (PEG)-mediated fusion with two immortal cell lines, Epithelioma papulosum cyprinid (EPC) and Spodoptera frugiperda pupal ovarian cells (Sf9), fused with Penaeus monodon haemocytes. The immortal cell lines were biochemically blocked with actinomycin D and puromycin before fusion occurred. A total of 78 hybrid clones were created. The methods used to confirm the presence of P. monodon genes and proteins in the hybrid cells did not detect crustacean components, nor was any viral amplification detected by real-time PCR after hybrid cells were inoculated with two P. monodon parvoviruses, Penaeus merguiensis densovirus and infectious hypodermal and haematopoietic necrosis virus. These results suggest although the creation of the hybrid cells appeared successful, the cell lines lacked key crustacean cell components required for their use as an in vitro system for virus replication.