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White spot syndrome virus (WSSV) is the most devastating pathogen found in shrimp aquaculture. The lack of certified continuous/established cell lines from penaeid shrimp restricts in vitro studies on the viruses to bring out effective prophylactic and therapeutic measures. In this context, a novel hybrid cell line named, PmLyO-Sf9, consisting of shrimp and Sf9 genomes has been established and employed to study WSSV susceptibility and multiplication. The hybrid cells were exposed to the shrimp virus WSSV and cytopathic effects (CPE) such as (a) enlargement of cells, (b) cessation cell division, (c) granulation of cytoplasm, (d) rounding off of cells, shortening and disappearance of tail-like structures and (e) detachment from the flask. Expression of immediate early genes such as ie 1, dnapol, rr1, tk-tmk, and pk 1could be confirmed indicating that viral DNA replication in the PmLyO-Sf9 took place followed by the expression of late genes such as VP-28, VP-26, VP-15 and VP-19. Electron micrograph of WSSV infected cells demonstrated marginated dense zones in the nucleus with clumped chromatin, and the mid zone with virus-like particles. However, neither discrete virus particles nor the culture supernatant having infectivity could be observed suggesting that virions were not getting formed in the cells. This is the first report of the susceptibility of PmLyO-Sf9 to WSSV, and the 'PmLyO-Sf9 - WSSV Complex' formed, defined as the infected status of PmLyO-Sf9 with WSSV, could be of use for unraveling at molecular level the mechanism of viral entry, replication impediments and cellular apoptosis., (Copyright © 2020. Published by Elsevier Inc.)

The White Spot Syndrome Virus (WSSV) has a large circular double-stranded DNA genome of around 300kb and it replicates in the nucleus of the host cells. The machinery of how the viral DNA is packaged has been remained unclear. VP15, a highly basic protein, is one of the major capsid proteins found in the virus. Previously, it was shown to be a DNA binding protein and was hypothesized to participate in the viral DNA packaging process. Using Atomic Force Microscopy imaging, we show that the viral DNA is associated with a (or more) capsid proteins. The organized viral DNA qualitatively resembles the conformations of VP15 induced DNA condensates in vitro. Furthermore, single-DNA manipulation experiments revealed that VP15 is able to condense single DNA against forces of a few pico Newtons. Our results suggest that VP15 may aid in the viral DNA packaging process by directly condensing DNA., (Copyright © 2010 Elsevier Inc. All rights reserved.)

White spot syndrome virus is one of the most serious viral pathogens causing huge mortality in shrimp farming. Here we report characterization of WSV010, a novel structural protein identified by our recent shotgun proteomics study. Its ORF contains 294 nucleotides encoding 97 amino acids. Transcription analysis using RT-PCR showed that wsv010 is a late gene. Localization analyses by Western blot and immunoelectron microscopy demonstrated that WSV010 is a viral envelope protein. Furthermore, the pull-down assay revealed that WSV010 could interact with VP24, which is a major envelope protein. Since WSV010 lacks a transmembrane domain, these results suggest that WSV010 may anchor to the envelope through interaction with VP24. Previous studies indicated that VP24 could also interact with VP28 and VP26. Therefore, we propose that VP24 may act as a linker protein to associate these envelope proteins together to form a complex, which may play an important role in viral morphogenesis and viral infection.

Based on a combination of SDS-PAGE and mass spectrometry, a protein with an apparent molecular mass of 31 kDa (termed as VP31) was identified from purified shrimp white spot syndrome virus (WSSV) envelope fraction. The resulting amino acid (aa) sequence matched an open reading frame (WSV340) of the WSSV genome. This ORF contained 783 nucleotides (nt), encoding 261 aa. A fragment of WSV340 was expressed in Escherichia coli as a glutathione S-transferase (GST) fusion protein with a 6His-tag, and then specific antibody was raised. Western blot analysis and the immunoelectron microscope method (IEM) confirmed that VP31 was present exclusively in the viral envelope fraction. The neutralization experiment suggested that VP31 might play an important role in WSSV infectivity.