Development and characterization of a new gill cell line from air breathing fish Channa striatus (Bloch 1793) and its application in toxicology: direct comparison to the acute fish toxicity.

A new cell line, Channa striatus gill (CSG), derived from the gill tissue of murrel, was established and characterized. The CSG cell line was maintained in Leibovitz's L-15 supplemented with 10% fetal bovine serum and has been subcultured more than 92 times. This cell line was able to grow in a range of temperatures from 22 to 32°C with optimal growth at 28°C. The plating efficiency was very high (52.21%) and doubling time was approximately 37h. The gill cell line was cryopreserved at different passage levels and revived successfully with 85% survival. Polymerase chain reaction amplification of mitochondrial 16S rRNA using primer specific to C. striatus confirmed the origin of this cell line from murrel. The cell line was further characterized by immunocytochemical analysis, chromosome number, transfection and mycoplasma detection. The cytotoxicity of endosulfan was assessed in CSG cell line using apoptosis assay, comet assay, mitochondrial alteration and five other endpoints such as Rhodamine 123 uptake, 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, neutral red assay, Alamar Blue assay and Methylene Blue protein assay. Acute toxicity study on fish was conducted by exposing murrel for 96h to endosulfan under static conditions. Statistical analysis revealed good correlation with r(2)=0.972-0.997 among the five endpoints. Linear correlations between the in vivo lethal concentration 50 (LC50) and each in vitro effective concentration 50 (EC50) were highly significant. The present study highlights the development of a new gill cell line from an air breathing fish that could be used as an alternative in vitro tools for studying pesticide toxicity in fish.