Species-independent bioassay for sensitive quantification of antiviral type I interferons

Abstract

BackgroundStudies of the host response to infection often require quantitative measurement of the antiviral type I interferons (IFN-α/β) in biological samples. The amount of IFN is either determined via its ability to suppress a sensitive indicator virus, by an IFN-responding reporter cell line, or by ELISA. These assays however are either time-consuming and lack convenient readouts, or they are rather insensitive and restricted to IFN from a particular host species.ResultsAn IFN-sensitive, Renilla luciferase-expressing Rift Valley fever virus (RVFV-Ren) was generated using reverse genetics. Human, murine and avian cells were tested for their susceptibility to RVFV-Ren after treatment with species-specific IFNs. RVFV-Ren was able to infect cells of all three species, and IFN-mediated inhibition of viral reporter activity occurred in a dose-dependent manner. The sensitivity limit was found to be 1 U/ml IFN, and comparison with a standard curve allowed to determine the activity of an unknown sample.ConclusionsRVFV-Ren replicates in cells of several species and is highly sensitive to pre-treatment with IFN. These properties allowed the development of a rapid, sensitive, and species-independent antiviral assay with a convenient luciferase-based readout.

Orthobunyaviruses and innate immunity induction: alieNSs vs. PredatoRRs

“…Results obtained for other orthobunyaviruses confirmed the conserved role of NSs as a virulence factor and suppressor of IFN induction (Blakqori et al, 2007;Elliott et al, 2013;Kraatz et al, 2015;Ogawa et al, 2007;Varela et al, 2013). Hence, orthobunyaviruses lacking NSs expression are convenient tools to trigger IFN induction in vitro and in vivo (Delhaye et al, 2006;Kallfass et al, 2012;Kuri et al, 2010), and are severely growth restricted in IFN-competent cells and animals (Barry et al, 2014;Blakqori et al, 2007;Bridgen et al, 2001;Kraatz et al, 2015;Ogawa et al, 2007;van Knippenberg et al, 2010;Varela et al, 2013;Weber et al, 2002). Such virus mutants are therefore prime candidates for attenuated live vaccines, as has been demonstrated for SBV (Kraatz et al, 2015).…”

Incoming RNA Virus Nucleocapsids Containing a 5′-Triphosphorylated Genome Activate RIG-I and Antiviral Signaling

“…D.Mel-2 cells (Gibco) were cultivated in Spodopan (Pan Biotech) at 28°C with no additional CO 2 . VSV, SFV, FLUAVΔNS1 (Garcia-Sastre et al, 1998), RVFVΔNSs::GFP (Habjan et al, 2008b; Kuri et al, 2010), RVFVΔNSs::REN (Habjan et al, 2008b; Kuri et al, 2010), and LACVdelNSs (Blakqori et al, 2007) were propagated on Vero cells. Plasmids pI.18_RVFV_N, pI.18_RVFV_L, pI.18_RVFV_M, and pHH21_RVFV_vMGFP were described previously (Habjan et al, 2008b; Habjan et al, 2009a).…”

Absence of a robust innate immune response in rat neurons facilitates persistent infection of Borna disease virus in neuronal tissue

“…Determination of IFN levels in cell culture medium were carried out as described previously [20]. In brief, rat C6 cells [21] were first seeded in 96-well plate and then incubated with known concentrations of rat IFN (Lee Biomolecular Research Inc., CA) or culture supernatant of the hippocampal slice culture for 24 h. Cells were then infected with VSV-GFP in DMEM containing 0.3 % bovine serum albumin (BSA) for 15 h. The GFP signal intensities were determined by a fluorescence reader (TECAN, Switzerland).…”