Overcoming host restrictions to enable continuous passaging of human noroviruses in human intestinal enteroids

PubMed: TBD

Authors

Gurpreet Kaur, Sue E. Crawford, Sara Javornik Cregeen, Anil Surathu, B. Vijayalakshmi Ayyar, Carmen V. Apostol, Hoa Nguyen Phuc, Khalil Ettayebi, Aaya Boussattach, Xi-Lei Zeng, Harsha Doddapaneni, Donna M. Muzny, Cristian Coarfa, B.V. Venkataram Prasad, Robert L. Atmar, Sasirekha Ramani, Mary K. Estes

Abstract

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 highly upregulated 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 with passaging 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 or immune evasion. This breakthrough in passaging provides new insight into HuNoV-host interactions, establishes a scalable in vitro system for virus propagation, and opens avenues for structural, biochemical, and therapeutic studies with virus stocks.

Code

This repo contains code and files needed to generate Figure 8 and Supplementary figures 6 & 7: Genetic changes during serial passages of GII.3 HuNoV in HIEs reveal one consensus change and dynamic variant shifts.