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Background Enteroviruses are non-enveloped, single-stranded positive-sense RNA viruses belonging to the family Picornaviridae. Enterovirus A71 (EV-A71) has caused recurrent outbreaks of hand, foot, and mouth disease especially among children in Asia. Some patients develop severe complications, such as meningitis, encephalitis, myocarditis, and pulmonary edema. A major hurdle for the development of antivirals for EV-A71 infection is the lack of robust antiviral platforms that closely mimic the in vivo setting. Organoids are laboratory-adapted miniaturized organs with preserved three-dimensional micro-anatomical architecture. In recent years, organoid cultures have been increasingly used for studying the pathogenesis of and evaluating antiviral treatment options for viral infections. In this study, we developed human intestinal organoids as a robust platform for evaluating antiviral options for EV-A71. Methods An epidemic strain of EV-A71 isolated from a patient with laboratory-confirmed EV-A71 infection was used. We compared the performance of multiple antiviral evaluation assays (virus yield reduction, plaque reduction, and cell protection assays) between human intestinal organoids and Caco-2 cells, using itraconazole (an antifungal previously shown to exhibit potent anti-enteroviral effects) and DMSO as positive and negative controls, respectively. Results The antiviral effect of itraconazole was comparable between human intestinal organoids and Caco-2 cells in the virus yield reduction and plaque reduction assays. In the cell protection assay, Caco-2 cells failed to demonstrate significant differences between the itraconazole-treated and DMSO-treated groups. In contrast, cell protection effects were easily observed and quantified in human intestinal organoids. Moreover, the human intestinal organoids allowed the characterization of the different cell types affected in EV-A71 infection with or without itraconazole treatment. Conclusion Human intestinal organoids support the replication of EV-A71 and provides a robust platform for antiviral evaluation for EV-A71 infection. Disclosures All authors: No reported disclosures.

2642. Development of Human Intestinal Organoids as an Antiviral Evaluation Platform for Enteroviruses