Preparation of Polyclonal Antiserum to the Recombinant TiLV-S8 Protein and Its Application in the Detection of Naturally Tilapia Lake Virus (TiLV) Infected Tilapia

Abstract

Tilapia lake virus (TiLV) is classified as a negative-sense, single-stranded RNA virus in the family Amnoonviridae. It is an enveloped virus with 10 genomic RNA segments, each coding for a protein. TiLV causes disease in tilapia, and outbreaks can lead to significant economic losses for the tilapia aquaculture industry. In this study, the gene encoding the segment 8 protein of TiLV was cloned into the expression vector pET15-b and then transformed into Escherichia coli strain BL21. After induction, the recombinant TiLV-S8 protein (rTiLV-S8), with a molecular mass of 20 kDa, was expressed, purified, and used to immunize mice. The mouse antiserum against rTiLV-S8 protein demonstrated specific immunoreactivity for the viral protein, approximately 19 kDa in TiLV-infected fish tissues, as determined by Western blotting. According to the results of the dot blotting assay, the antiserum was about 80 times less sensitive than one-step RT-PCR in detecting TiLV in homogenates of infected fish samples and showed no cross-reaction with uninfected fish tissues, other common fish viruses, or prevalent bacterial species found in aquatic animals. Furthermore, this polyclonal antiserum could be employed to identify TiLV-infected fish in the field using dot blotting assay, and the results can be confirmed by immunohistochemistry.