Development and characterization of novel cell lines from kidney and eye of rainbow trout, Oncorhynchus mykiss for virological studies

Abstract

Two novel cell lines were developed from eye and kidney of rainbow trout, Oncorhynchus mykiss in addition to another cell line from spleen. All these cell lines were characterized and cryopreserved successfully at different passage levels. The cells of eye, kidney and spleen were sub-cultured continuously at an interval of 2–3 days and attained the passage level of 112, 106 and 112, respectively over a period of 2 years. These cell lines consisted of both fibroblastic and epithelial like cells, and showed the presence of pancytokeratin and fibronectin by immunofluorescence. Maximum growth of cells was observed at 16 °C and 15% concentration of FBS. Amplification and sequence analysis of COI of O. mykiss confirmed the origin of these cell lines from O. mykiss. These cell lines were found to be highly susceptible to fish nodavirus (FNV) and tilapia lake virus (TiLV), and resistant to Cyprinid herpesvirus 2 (CyHV-2). The RT-PCR assay confirmed the infection of TiLV and FNV in infected cell lines. The FNV infection was further confirmed by Western blot and ELISA. The titre value of FNV in infected RTE, RTS and RTK cells gradually increased from 103.2, 102.9, 103.3 TCID50 ml−1 at 2 d.p.i, to 108.2, 107.9, 108.8 TCID50 ml−1 at 10 d.p.i. respectively. Attempt was made to use these cell lines for virus isolation from disease suspected fish samples collected from different districts of Assam state, India.