Establishment and characterization of two cell lines from bluefin trevally Caranx melampygus

Abstract

Bluefin trevally Caranx melampygus Cuvier is a popular game fish and highly valued sea food with potential importance for aquaculture. To help establish this marine animal as an important aquacultural species in Hawaii and the Pacific and develop in vitro cell culture systems for long-term management and control of potential viral diseases 2 cell lines were established from body muscle (bluefin trevally muscles, BTMS) and fins (bluefin trevally fins, BTF). Primary culture of these cells was conducted at 25°C using L-15 medium supplemented with 20% fetal bovine serum and various antibiotics. These cells have been serially subcultured 37 to 41 times since their initiation in June 2002. Growth of the bluefin trevally cells was serum-dependent at the time of the experiments and their plating efficiencies ranged from 11 to 28.2%. Comparative analysis showed that these bluefin trevally cells grew equally well in the media L-15 (Leibovitz medium), RPMI 1640, M199 and MEM (minimum essential medium), which are commonly used for cell cultures derived from aquatic animals and mammalian species. Examination of the early passage cells stored at -196°C revealed a large percent (nearly 98%) of cell viability following a 6 mo storage in liquid nitrogen. Karyotyping analysis indicated that these bluefin trevally derived cell lines remained diploid with a chromosome count of 48 at passage 7 and 12. These 2 cell lines shared a same pattern of viral susceptibility and they were sensitive to infectious hematopoietic necrosis virus (IHNV), infectious pancreatic necrosis (IPN), spring viremia carp virus (SVCV), viral hemorrhagic septicemia virus (VHSV), and snakehead rhabdovirus (SHRV) but refractory to channel catfish virus (CCV) infection. These newly established cell lines are currently being used to facilitate the diagnosis of viral disease affecting marine fish aquaculture in Hawaii, and will be available for future isolation and study of bluefin trevally fish viruses.

Establishment, characterization, and viral susceptibility of two cell lines derived from goldfish Carassius auratus muscle and swim bladder

“…This method was adapted accordingly from previous cell line establishment efforts (Lu et al 1999, Zhao & Lu 2006. Each cell line was originally seeded in 25 cm 2 tissue culture flasks and later passed into 75 cm 2 tissue culture flasks for cultivation (Corning Life Sciences).…”

“…This method was adapted accordingly from previous cell line establishment efforts (Lu et al 1999, Zhao & Lu 2006 ) were performed using a hemocytometer. Analogous procedures were performed for the effects of various concentrations of FBS (0, 5, 10, 20%) and the different cell culture media (L-15, RPMI 1640, Minimum Essential Medium and Medium 199) on cell growth over 10 d.…”

Spring viraemia of carp virus: recent advances

“…SVCV has been reported to infect and multiply in various fish cell lines, including epithelioma papulosum cyprinid cells from carp (Gotesman et al, 2015); skin cells from Ussuri catfish (Pseudobargus ussuriensis) and red-spotted grouper (Epinephelus akaara) (Lei et al, 2014;Ou et al, 2012); heart cells from giant grouper (Epinephelus lanceolatus) (Guo et al, 2015); haploid embryonic cells from medaka (Oryzias latipes) (Yuan et al, 2013); muscle and fin cells from bluefin trevally (Caranx melampygus) (Zhao & Lu, 2006); swim bladder, fin and snout cells from grass carp (Lu et al, 1990); and muscle, heart and swim bladder cells from snakehead (Channa striatus) (Zhao et al, 2004). SVCV propagation in these cells has been analysed by RT-PCR, electron microscopy, immunofluorescence assay and 50% TCID assays.…”

Development and characterization of cell culture systems from Puntius (Tor) chelynoides (McClelland)

“…This suggests the utility of COI sequence as a valid and universal marker for species identification of established fish cell lines. Other alternatives such as 16S ribosomal RNA gene sequence has also been used to confirm the origin of muscle and fin cell lines of bluefin trevally (Zhao and Lu, 2006). Identity of PCE cell line was confirmed by COI and 16S rRNA genes amplification.…”