Modification of H1N1 Influenza Luciferase Reporter Viruses Using StopGo Translation and/or Mouse-Adapted Mutations

Area of Science:

• Virology
• Molecular Biology
• Biotechnology

Background:

• Reporter viruses are essential for studying viral infections and developing antiviral therapies.
• Previous influenza reporter viruses showed growth defects and required mouse adaptation for improved performance.

Purpose of the Study:

• To investigate if a growth defect in a previously developed influenza reporter virus (rTN09-PA-Nluc) was due to PA-Nluc protein fusion.
• To engineer a new reporter virus (rTN09-PA-Nluc/SG) utilizing StopGo translation to produce separate PA and Nluc proteins.

Main Methods:

• Generation of the rTN09-PA-Nluc/SG reporter virus with StopGo translation.
• Comparison of protein expression, replication, and neutralization assays in vitro between rTN09-PA-Nluc and rTN09-PA-Nluc/SG.
• Assessment of reporter virus virulence and imaging capabilities in mice.

Main Results:

• The StopGo translation virus (rTN09-PA-Nluc/SG) exhibited enhanced protein expression and replication in cell culture compared to the fusion virus.
• Bioluminescence-based virus neutralization assays using the StopGo virus yielded results in 24 hours, significantly faster than standard methods.
• The StopGo virus remained attenuated in mice, requiring mouse-adaptive mutations for full virulence and efficient in vivo imaging.

Conclusions:

• Incorporating StopGo translation into influenza reporter viruses significantly benefits in vitro assays, enabling rapid results.
• While StopGo translation improves in vitro performance, mouse-adapted mutations are crucial for optimal virulence and imaging in vivo.
• This study highlights the distinct requirements for reporter virus efficacy in cell culture versus animal models.