Abstract
Amphiprion ocellaris (ocellaris clownfish) is one of the most commercially important marine ornamental fish. A cell line designated as OCF was developed for the first time from the caudal fin of this fish species. The cell line was maintained in Leibovitz’s—15 medium supplemented with 15% FBS (Fetal Bovine Serum) and was successfully subcultured up to 34 passages. The cell line was authenticated by sequencing mitochondrial cytochrome C oxidase subunit I (COI) and 16S rRNA genes. The growth rate of the OCF cell line was maximum in medium containing 20% FBS and 1% of 0.2 M NaCl at 28 °C. Chromosome analysis revealed 48 diploid chromosomes. The OCF cell line was transfected with the pMaxGFP plasmid vector with 7% efficiency and GFP expression was observed. The OCF cell line was used for testing nervous necrosis virus (NNV) susceptibility. Cytopathic effect (CPE) was observed in terms of plaque formation after virus inoculation. Nested PCR confirmed the susceptibility of the OCF cell line to NNV. The cell line was successfully cryopreserved by a slow freezing procedure at − 80 °C with a revival efficiency of 70–75%. The study revealed that the OCF cell line would be useful for virological studies. In addition, the cell line would play an important role as an in vitro tool for carrying out toxicological and biotechnological studies.
Highlights
Amphiprion ocellaris is one of the most commercially important marine ornamental fish
Cell growth was observed at different concentrations of Fetal Bovine Serum (FBS), i.e., 5, 10, 15, and 20% to find out optimum FBS concentration
Ocellaris clownfish fin (OCF) cells revealed poor growth at 5% of FBS concentration whereas comparatively better growth was observed at 15% (Table S1, Fig S1), but maximum growth of the cells was observed with 20% FBS concentrations (Fig. 2)
Summary
Amphiprion ocellaris (ocellaris clownfish) is one of the most commercially important marine ornamental fish. Fish cells have an advantage over mammalian or avian cells with minimal maintenance requirement, replicate within a broad range of incubation temperatures, and a flexible culture p rogram[3] These distinctive features of fish cells make them useful tools for a variety of biological requisitions in life science. The catch of the clownfish has reduced dramatically in the last few years because of over-exploitation in response to its increasing demand, popularity, and worsening of its natural habitats These scrutinize have led to the captive breeding of these marine ornamental fish, for conservation as well as commercial purposes[11,12]. To undertake in vitro studies on the viruses infecting the species, a suitable cell line is not available In this background, a cell line was developed for the first time using the caudal fin of marine ornamental fish, Amphiprion ocellaris. The mass mortality was observed in hatchery-reared larvae of clownfish due to NNV or b etanodavirus[59]
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