Gurpreet Kaur, Sue Crawford, Sara Javornik Cregeen, Anil Surathu, B. Vijayalakshmi Ayyar, Carmen V. Apostol, Hoa Nguyen Phuc, Khalil Ettayebi, Aaya Boussattach, Xi-Lei Zeng, Sarah E. Blutt, HarshaVardhan Doddapaneni, Donna M. Muzny, Cristian Coarfa, Ramakrishnan Anish, B. V. Venkataram Prasad, Robert L. Atmar, Sasirekha Ramani, Mary K. Estes
The establishment of human intestinal enteroids (HIEs) as a model for human norovirus (HuNoV) replication has been transformative for studying this leading cause of gastroenteritis. However, indefinite passaging of HuNoVs in HIEs remained a challenge, necessitating the use of patient stool samples as viral inocula. Using RNA-seq, we identified CXCL10, CXCL11, and CCL5 as up-regulated chemokines, suggesting their potential as host restriction factors. TAK-779, a CXCR3/CCR5/CCR2 antagonist, enhanced GII.3 HuNoV replication and viral spread in a dose- and time-dependent manner, enabling successful passaging of GII.3 HuNoV in two different HIE lines and generation of viral stocks. Sequencing passaged virus revealed one consensus change in the major capsid protein and several dynamic adaptations, suggesting emergence of variants. TAK-779 also enhanced replication of GI.1 and GII.17 strains, but not GII.4, suggesting strain-specific host interactions. This breakthrough in passaging provides insight into HuNoV-host interactions, establishes a scalable in vitro system for virus propagation, and opens avenues for structural, biochemical, and therapeutic studies.