High-Throughput Screening of Antiviral Agents Using the Fluorescence-Based Local Lesion Inspection Assay.

Area of Science:

• Plant pathology
• Virology
• Drug discovery

Background:

• Viral diseases pose significant threats to agriculture and human health.
• Developing effective antiviral agents requires efficient screening methods.
• Understanding the mechanism of action (replication vs. movement inhibition) is crucial for drug development.

Purpose of the Study:

• To develop and present a high-throughput screening system for identifying antiviral agents.
• To establish a method for differentiating the mechanism of action of identified antiviral agents.
• To provide a detailed protocol for the fluorescence-based local lesion inspection assay (FLLIA).

Main Methods:

• Mechanical inoculation of plant leaves with a fluorescent protein-expressing engineered virus.
• Application of test chemicals to one half of the leaf and buffer to the other half.
• Imaging and quantification of viral infection lesion number, size, and fluorescence intensity.

Main Results:

• The FLLIA system enables high-throughput screening of potential antiviral compounds.
• Reduced lesion number/fluorescence intensity indicates replication inhibition.
• Reduced lesion size indicates movement inhibition.

Conclusions:

• FLLIA is a simple and efficient method for identifying antiviral agents.
• The assay effectively differentiates between replication and movement inhibition mechanisms.
• This system aids in the discovery and characterization of novel antiviral therapies.