Development and Application of MLB Human Astrovirus Reverse Genetics Clones and Replicons.

Human astroviruses pose a significant public health threat, especially to children, the elderly, and immunocompromised individuals. Nevertheless, these viruses remain largely understudied, with no approved antivirals or vaccines. This protocol focuses on leveraging reverse genetics (RG) and replicon systems to unravel the biology of MLB genotypes, a key group of neurotropic astroviruses. Using reverse genetics and replicon systems, we identified critical genetic deletions linked to viral attenuation and neurotropism, pushing forward vaccine development. We also uncovered novel replication mechanisms involving ER membrane interactions, opening doors to new antiviral targets. Reverse genetics and replicon systems are essential for advancing our understanding of astrovirus biology, identifying virulence factors, and developing effective treatments and vaccines to combat their growing public health impact. Key features • Provides a basic understanding of the molecular biology of MLB astroviruses, aiding in addressing open questions related to virus evolution, replication, and pathogenesis. • Facilitates the development of novel therapeutics and vaccines. • Enables rapid testing of antiviral drugs against MLB astroviruses.