Real-time monitoring of Equid herpesviruses infectivity in equine dermal cell based on impedance measurements
Résumé
In virology, different conventional methods are commonly used for virus titration and infectivity studies. Most of them are based on direct examination of viral cytopathic effect, which is time consuming, tedious and requires an endpoint reading. The RTCA xCELLigence system (ACEA Biosciences) has been developed to follow cellular events and their dynamics in real time using a micro-electrical biosensor technology. The aim of this study was to evaluate the feasibility of using this biosensor technology to measure the effect of different equid herpesviruses on equine dermal cell cultures, with or without antiviral acyclovir treatment. Cells were infected with EHV-1 (VR700), EHV-4 (VR2230), EHV-2 (VR701) or EHV-3 (VR352). The viral effects were monitored by 3 different methods: impedance measurements (RTCA), cytopathic effects recording (microscopy) and viral load quantification (qPCR). The RTCA technology showed a dose dependent drop of the cellular index induced by specific impedance variations measured during growth of each virus tested. These results correlated with the cytopathic effect induced by the different herpesvirus species, as well as viral loads measured by qPCR. For example, after 3 days of incubation with EHV-1, RTCA results showed an increase of the cellular index from 0% (infection without antiviral) to 45% and 100% in presence of Acyclovir (10μl/mL and 100 μg/mL, respectively). These results are correlated with the decreased of viral load measured in the presence of Acyclovir: 1010 genome copies/mL with EHV-1 only, 106 copies/mL with EHV-1 + Acyclovir 10 μg/ mL and 105 copies/mL with EHV-1 + Acyclovir 100 μg/ mL. In conclusion, the cellular impedance monitoring system is a new real-time method to monitor cell growth and viral proliferation in cell cultures. This technology is well adapted for high-throughput screening of antiviral molecules. In fact, the xCELLigence system enables continuous impedance measurement and quantification of cell adhesion, proliferation, cell death and detachment during viral infection.