Recombinant Lloviu virus as a tool to study viral replication and host responses

Area of Science:

• Virology
• Molecular Biology
• Pathogen Discovery

Background:

• Next-generation sequencing identifies numerous RNA viruses in animal hosts with zoonotic potential.
• Many of these viruses, like Lloviu virus (LLOV), remain uncultured and understudied, hindering research.
• Reverse genetics systems are crucial for rescuing viruses but often require complete viral genome sequences.

Purpose of the Study:

• To generate infectious Lloviu virus (LLOV) from rescued clones.
• To identify cis-acting elements essential for LLOV replication.
• To investigate the tropism and cellular response to LLOV infection.

Main Methods:

• Utilized minigenome systems to complement missing LLOV genomic ends.
• Generated recombinant full-length LLOV clones for virus rescue.
• Employed electron microscopy to characterize rescued virus morphology and inclusions.
• Performed cell tropism assays on known Ebola virus target cells and pneumocytes.
• Tested antiviral efficacy against the rescued LLOV.

Main Results:

• Successfully rescued infectious recombinant LLOV (rLLOV) by complementing genomic ends and identifying cis-acting elements.
• rLLOV exhibited filamentous virions and cytoplasmic inclusions, characteristic of filoviruses.
• Macrophages, hepatocytes, and pneumocytes were identified as permissive to rLLOV infection.
• rLLOV infection did not induce inflammatory responses in human macrophages, unlike Ebola virus.
• Antivirals targeting various replication stages were tested using rLLOV.

Conclusions:

• Rescue of uncultured viruses like LLOV is feasible and provides a vital tool for studying emerging pathogens.
• rLLOV serves as a model for investigating filovirus replication, tropism, and antiviral strategies.
• Understanding LLOV's cellular interactions and host responses is crucial for assessing its zoonotic risk.
• This work enhances pandemic preparedness by enabling research on dangerous, previously uncultured viruses.