Visualizing infection by single positive-sense RNA viruses using virus infection real-time imaging (VIRIM).

Area of Science:

• Virology
• Cell Biology
• Molecular Biology

Background:

• Conventional assays for viral infection often average cell populations, masking critical cell-to-cell variability.
• Existing methods typically require cell fixation or lysis, preventing real-time observation of infection dynamics.

Purpose of the Study:

• To develop a novel real-time, single-cell assay for tracking viral RNA translation.
• To enable the study of early viral infection events and virus-host interactions at the single-molecule level.
• To investigate the origins and consequences of heterogeneity during viral infections.

Main Methods:

• Development of virus infection real-time imaging (VIRIM), a live-cell imaging technique.
• Visualization of individual viral RNA translation in real-time using single-molecule detection.
• Detailed protocols for virus and reporter cell line generation, time-lapse imaging, and data analysis.

Main Results:

• VIRIM allows visualization of viral RNA translation at the single-molecule level in live cells.
• The assay captures the earliest stages of viral infection, even with low viral RNA and protein levels.
• Enables detailed examination of cell-to-cell heterogeneity in viral infection progression.

Conclusions:

• VIRIM provides unprecedented insight into the dynamics of viral infection at the single-cell level.
• This method is crucial for understanding virus-host interactions and the heterogeneity of infection.
• The VIRIM assay offers a powerful tool for virology research, requiring 2-5 days for completion.