A New Cell Line from the Brain of Red Hybrid Tilapia (Oreochromis spp.) for Tilapia Lake Virus Propagation

Author:

Mohamad Aslah1ORCID,Khemthong Matepiya1,Trongwongsa Pirada1,Lertwanakarn Tuchakorn2ORCID,Setthawong Piyathip2,Surachetpong Win13ORCID

Affiliation:

1. Department of Veterinary Microbiology and Immunology, Faculty of Veterinary Medicine, Kasetsart University, Bangkok 10900, Thailand

2. Department of Physiology, Faculty of Veterinary Medicine, Kasetsart University, Bangkok 10900, Thailand

3. Laboratory of Biotechnology, Chulabhorn Research Institute, Bangkok 10210, Thailand

Abstract

Tilapia lake virus (TiLV) presents a substantial threat to global tilapia production. Despite the development of numerous cell lines for TiLV isolation and propagation, none have been specifically derived from red hybrid tilapia (Oreochromis spp.). In this study, we successfully established a new cell line, RHTiB, from the red hybrid tilapia brain. RHTiB cells were cultured for 1.5 years through over 50 passages and demonstrated optimal growth at 25 °C in Leibovitz-15 medium supplemented with 10% fetal bovine serum at pH 7.4. Morphologically, RHTiB cells displayed a fibroblast-like appearance, and cytochrome oxidase I gene sequencing confirmed their origin from Oreochromis spp. Mycoplasma contamination testing yielded negative results. The revival rate of the cells post-cryopreservation was observed to be between 75 and 80% after 30 days. Chromosomal analysis at the 25th passage revealed a diploid count of 22 pairs (2n = 44). While no visible cytopathic effects were observed, both immunofluorescence microscopy and RT-qPCR analysis demonstrated successful TiLV propagation in the RHTiB cell line, with a maximum TiLV concentration of 107.82 ± 0.22 viral copies/400 ng cDNA after 9 days of incubation. The establishment of this species-specific cell line represents a valuable advancement in the diagnostic and isolation tools for viral diseases potentially impacting red hybrid tilapia.

Funder

Office of the Ministry of Higher Education, Science, Research and Innovation and the Thailand Science Research and Innovation

Office of the National Economic and Social Development Council, the Office of the Prime Minister

Kasetsart University Research and Development Institute

Publisher

MDPI AG

Reference45 articles.

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