An improved reverse genetics system for rotavirus vaccine strain LLR using five plasmid vectors

Area of Science:

• Virology
• Molecular Biology
• Vaccinology

Background:

• Rotaviruses (RVs) are a major cause of severe gastroenteritis in young children.
• Species A rotaviruses have an 11-segmented double-stranded RNA genome.
• Previous work established a method to recover RV strain LLR using 11 plasmids.

Purpose of the Study:

• To improve the reverse genetics system for rotavirus vaccine strain LLR.
• To enhance the efficiency of rotavirus rescue and recombinant virus generation.
• To facilitate studies on rotavirus biology and clinical applications.

Main Methods:

• Developed a novel reverse genetics system by combining transcriptional cassettes (two or three) into single plasmids.
• Utilized the improved system to rescue rotavirus vaccine strain LLR.
• Generated a recombinant LLR strain stably expressing NLuc using a five-plasmid system.

Main Results:

• Achieved a significant enhancement in rescue efficiency from 66.7% (8/12) to 91.7% (11/12).
• Successfully rescued a recombinant LLR strain stably expressing NLuc.
• Demonstrated the improved system's effectiveness for generating genetically modified rotaviruses.

Conclusions:

• The optimized reverse genetics system substantially increases rotavirus rescue efficiency.
• This improved system is valuable for generating recombinant rotaviruses for research and potential clinical use.
• Advancements in rotavirus reverse genetics will accelerate studies on RV pathogenesis and vaccine development.