Development of high-throughput screening viral titration assay: Proof of concept through two surrogate viruses of human pathogens

The precise determination of viral titers in virological studies is a critical step to assess the infectious viral concentration of a sample. Although conventional titration methods, such as endpoint dilution or plaque forming units are the gold standards, their widespread use for screening experiments remains limited due to the time-consuming aspect and resource-intensive requirements. This study introduces a rapid and user-friendly high-throughput screening assay for evaluating viral titers. The colorimetric method used relies upon assessing virus-induced cytopathic effects by measuring the reduction of a tetrazolium reagent to formazan through cellular dehydrogenation within mitochondria. The resulting formazan quantity is correlated with the viral titer and can be easily quantified by a colorimetric measurement. In this perspective, this manuscript describes two case studies for the titration of the porcine respiratory coronavirus virus and bovine alpha herpesvirus 1, highlighting, respectively, a linear regime between 100 and 2000 TCID₅₀/ml and 500- <mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML"> <mml:mrow> <mml:msup><mml:mrow><mml:mn>10</mml:mn></mml:mrow> <mml:mrow><mml:mn>6</mml:mn></mml:mrow> </mml:msup> </mml:mrow> </mml:math> PFU/ml for rapid titration within these ranges. The proposed technique's advantages and drawbacks are discussed, along with potential applications such as drug screening and the assessment of viral survival on inert surfaces.