The system currently used at AGES detects the viral E- and -RdRp gene with internal control (EAV, Equine Arteritis Virus) according to the WHO recommendation: Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR (Victor M Corman et al., 2020) and is a "homebrew" system using primers, probes and controls from TIB-MolBiol (Berlin) and buffer/enzyme from Invitrogen SuperScriptTM III One-Step RT-PCR System (ThermoFischer).

In the first approach ("screening"), a 76 base pair long fragment of the viral E-gene and the internal control (EAV, extraction and PCR control) are detected. Positive samples in E-gene PCR are additionally tested with RdRp (100 base pairs) for confirmation. The second system, a "rapid test" with isothermal amplification from MIRAI GENOMICS uses the SmartAmp 2nd kit from RIKEN (Japan). The third test kit, ViroReal® Kit SARS-CoV-2 & SARS, uses the viral N gene as a target sequence and is produced by Ingenetix (Vienna). The three systems tested matched in 90 of the 94 patient samples tested. In two cases (samples 850500 and 850514), the confirmatory test was negative after positive E gene with RdRp (routine system AGES), but both MIRAI and Ingenetix were positive. Thereby, the "Cp values" were very high in isothermal amplification (75.25 and 82.32). Both MIRAI (850461) and Ingenetix (850502) gave positive results with negative E and RdRp genes.

Florian Gruber, MedUni Vienna Peter Hufnagl, AGES