Temporary protection of rainbow trout gill epithelial cells from infection with viral haemorrhagic septicaemia virus IVb

Abstract

The branchial epithelium is not only a primary route of entry for viral pathogens, but is also a site of viral replication and subsequent shedding may also occur from the gill epithelium. This study investigated the potential of agents known to stimulate innate immunity to protect rainbow trout epithelial cells (RTgill-W1) from infection with VHSV IVb. RTgill-W1 cells were pretreated with poly I:C, FuGENE(®) HD + poly I:C, lipopolysaccharide (LPS), LPS + poly I:C or heat-killed VHSV IVb and then infected with VHSV IVb 4 days later. Cytopathic effect (CPE) was determined at 2, 3, 4, 7 and 11 days post-infection. Virus in cells and supernatant was detected using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). All of the treatments delayed the onset of CPE (per cent of monolayer destruction), compared with untreated controls; however, killed VHSV or poly I:C combined with LPS was the most effective. Similarly, the detection of viral RNA in the supernatant was delayed, and the quantity was significantly (P < 0.05) reduced by all treatments with the exception of LPS alone (4 days). Unlike many of the other treatments, pretreatment of RTgill-W1 with heat-killed VHSV did not upregulate interferon 1, 2 or MX 1 gene expression.

Autophagy‐related genes in rainbow trout Oncorhynchus mykiss (Walbaum) gill epithelial cells and their role in nutrient restriction

“…The rainbow trout gill epithelial cell line RTgill-W1 (Bols et al, 1994) was used because our laboratory previously demonstrated that reduced viral haemorrhagic septicaemia virus (VHSV) IVb was present in this cell line after treatment with killed VHSV but that stimulation of many known anti-viral genes was not responsible (Al-Hussinee et al, 2016). As a prelude to understanding the potential role of autophagy in disease resistance, the purpose of this study was to expand the repertoire of rainbow trout Atg genes and their induction under nutrient restriction in vitro in RTgill-W1.…”

VHSV IVb infection and autophagy modulation in the rainbow trout gill epithelial cell line RTgill‐W1

“…However, the complexity of the interaction between host and pathogen has led to the demonstration that variable outcomes occur even when similar research models are employed (Garcia‐Valtanen et al., 2014; Liu et al., 2015). Previously, the rainbow trout epithelial cell line RTgill‐W1 was shown to be susceptible to VHSV IVb infection (Pham et al., 2013), but pretreatment of the cells with heat‐inactivated VHSV IVb reduced virus replication without increased expression of antiviral cytokines (Al‐Hussinee et al., 2016). Therefore, we investigated the role of autophagy in the RTgill‐W1 during infection with VHSV IVb.…”

“…A one‐step RT‐qPCR was used to detect the VHSV IVb viral RNA copy number in the cellular RNA extraction (Al‐Hussinee et al., 2016). The LightCycler 480 RNA Master Hydrolysis Probes (Roche) Kit was applied in the LightCycler 480 Instrument II System (Roche) according to the instruction of the manufacturer.…”

“…The rainbow trout gill epithelial cell line, RTgill‐W1, was found to be susceptible to VHSV IVb, while a monocyte/macrophage cell line (RTS11) was not (Pham, Lumsden, Tafalla, Dixon, & Bols, 2013). Pretreatment of rainbow trout gill epithelial cells (RTgill‐W1) with inactivated VHSV IVb reduced subsequent VHSV replication without upregulation of some key antiviral genes including mx1 , ifn1 and ifn2 (Al‐Hussinee et al., 2016).…”

Polarized Trout Epithelial Cells Regulate Transepithelial Electrical Resistance, Gene Expression, and the Phosphoproteome in Response to Viral Infection

“…The use of cell cultures such as the RTgill-W1 cell line developed from a primary culture of rainbow trout gill Oncorhynchus mykiss (Walbaum) ( 13 ) can be very useful to delineate specific molecular and cellular activation pathways. The RTgill-W1 cell line has been used to study the antiviral response ( 14 ), the effect of osmoregulatory hormones ( 15 ) and in ecotoxicological studies ( 16 ). In RTgill-W1 cells cultured under transwell conditions canonical features of normal epithelial function are observed such as pavement cells (PVCs) and microridges however are absent in flask culture.…”