Isolation and characterization of a novel strain (YH01) of Micropterus salmoides rhabdovirus and expression of its glycoprotein by the baculovirus expression system

Abstract

As one of the most important aquatic fish, Micropterus salmoides suffers lethal and epidemic disease caused by rhabdovirus at the juvenile stage. In this study, a new strain of M. salmoides rhabdovirus (MSRV) was isolated from Yuhang, Zhejiang Province, China, and named MSRV-YH01. The virus infected the grass carp ovary (GCO) cell line and displayed virion particles with atypical bullet shape, 300-500 nm in length and 100-200 nm in diameter under transmission electron microscopy. The complete genome sequence of this isolate was determined to include 11 526 nucleotides and to encode five classical structural proteins. The construction of the phylogenetic tree indicated that this new isolate is clustered into the Vesiculovirus genus and most closely related to the Siniperca chuatsi rhabdovirus. To explore the potential for a vaccine against MSRV, a glycoprotein (1-458 amino acid residues) of MSRV-YH01 was successfully amplified and cloned into the plasmid pFastBac1. The high-purity recombinant bacmid-glycoprotein was obtained from DH10Bac through screening and identification. Based on polymerase chain reaction (PCR), western blot, and immunofluorescence assay, recombinant virus, including the MSRV-YH01 glycoprotein gene, was produced by transfection of SF9 cells using the pFastBac1-gE2, and then repeatedly amplified to express the glycoprotein protein. We anticipate that this recombinant bacmid system could be used to challenge the silkworm and develop a corresponding oral vaccine for fish.

The recombinant subunit vaccine encapsulated by alginate‐chitosan microsphere enhances the immune effect against Micropterus salmoides rhabdovirus

“…Viruses (MSRV‐FJ985) were kept well in our laboratory. MSRV‐FJ985 was incubated on GCO (grass carp ovary) cells at 25°C in 5% CO 2 atmosphere in Medium 199 (Hyclone, USA) cell culture containing 10% foetal bovine serum (ZETA LIFE, USA), streptomycin 100 U/ml and penicillin 100 U/ml (Lyu et al, 2019; Yang et al, 2021). Virus titers were determined for challenge.…”

“…Rhabdovirus genomes generally encode (from 3′ to 5′) five major structural proteins including: nucleoprotein (N), phosphorylated protein (P), matrix protein (M), glycoprotein (G) and RNA polymerase protein (L), all of which play an important role in virus infection and replication (Dietzgen et al, 2017; Feng et al, 2021). Among the five core proteins, glycoproteins are the main structural proteins that form viral spike and widely used in anti‐viral vaccine construction (Lyu et al, 2019; Zhang, Li, Lin et al, 2018). Over the past decades, vaccines represent one of the most effective strategies to protect fish against viral diseases.…”

Revised Taxonomy of Rhabdoviruses Infecting Fish and Marine Mammals

“…Micropterus salmoides rhabdovirus (MSRV) was first discovered in juvenile mandarin fish collected in Guangdong Province, China, in 2011 [ 36 ]. A second sample (YH01; GenBank MK397811) was isolated from moribund largemouth bass ( Micropterus salmoides ) collected in Zhejiang Province, China, in 2017 [ 37 ]. A third isolate of the virus (FJ985; GenBank MT818233) was obtained from China in 2019.…”

Rapid visual detection of Micropterus salmoides rhabdovirus using recombinase polymerase amplification combined with lateral flow dipsticks

“…Unfortunately, a new viral disease impacted the largemouth bass aquaculture industry in Guangdong Province, China, in 2011, resulting in 200,000 fish deaths and substantial economic losses (Ma et al., 2013). Subsequently, rhabdovirus was confirmed as the pathogen and several rhabdovirus isolates were reported (Fu et al., 2017; Lyu et al., 2019; Ma et al., 2013).…”