Establishment and characterization of three cell lines derived from the rockfish grouper Epinephelus quoyanus: Use for transgenic studies and cytotoxicity testing

The establishment and partial characterization of continuous cell lines from Epinephelus quoyanus are described. Three new cell lines, designated as RGB, RGG and RGH, were established from brain, gill and heart tissue, respectively, of the rockfish grouper E. quoyanus. Since their initiation in December of 2006, these cell lines have been subcultured in Leibovitz's L-15 medium with 10% fetal bovine serum for more than 80 passages at 30 °C. The morphologies of RGB and RGG cells were epithelioid, while RGH was predominantly fibroblast-like. Propagation of these cell lines was serum-dependent, with a low plating efficiency (< 4%). Karyotyping analysis indicated that the modal chromosome number was 48 for each cell type. The RGB and RGH cells were susceptible to grouper nervous necrosis virus (GNNV) but only yielded titers of 104.8 and 104.6 TCID50 ml− 1, respectively. Moreover, RGB was susceptible to hard clam reovirus (HCRV) and yielded titers of 107.2 TCID50 ml− 1. All three cell lines were also sensitive to the extracellular products of Photobacterium damselae ssp. piscicida (Ph.d.p.) and produced significant fluorescent signals after transfection with pEGFP-C3. Analysis of mitochondrial cytochrome b gene sequences revealed 99% identity among the cell lines and E. quoyanus. Our results suggest that the three cell lines provide a valuable tool for transgenic studies and the investigation of the extracellular products of Ph.d.p.