Human enterochromaffin cells and apical-out intestinal organoids as models for human sapovirus infection

Human sapovirus is increasingly recognized as a significant cause of acute viral gastroenteritis, but in vitro studies have been limited due to the previous lack of cell models. In this study, we optimized two novel in vitro models for sapovirus infection: the human enteroendocrine cell (EEC) line GOT1 and apical-out 3D human enteroids. Using 31 sapovirus-positive fecal samples, we compared these models with the EEC-derived HuTu80, and human enteroid monolayers, both previously used for sapovirus infection. Among the analyzed samples, genotypes GI.1 (36%), GII.1 (32%), and GII.3 (26%) were identified, with GI.1 exhibiting the highest fecal viral load. We demonstrated that GOT1 cells supported replication of more of the samples containing GI.1 sapovirus (9 of 11) compared to HuTu80 (8 of 11) and with a generally higher replication fold change. Moreover, GOT1 cells were able to support the replication of several GII.1 (3 of 10) and GII.3 (5 of 8) samples, unlike HuTu80 cells that only supported replication of one sample with GII.3 and none of the GII.1 sapoviruses. Given that organoids are considered a more physiologically relevant model than transformed cell lines, we established a sapovirus infection model using human apical-out 3D enteroids. Compared to previously used enteroid monolayers, the apical-out model supported replication of a higher number of samples with GI.1 and with higher replication fold change. In conclusion, these findings provide valuable insights for future in vitro studies of sapovirus infection and replication, which may contribute to a better understanding of sapovirus cell tropism and pathogenesis. ImportanceUnderstanding sapovirus biology requires efficient, reliable, and physiologically relevant in vitro models. By systematically comparing four infection models using clinical samples from three common sapovirus genotypes, this study contributes to important information about differences in cellular susceptibility between these in vitro models. We identified the GOT1 cell line as an efficient model for sapovirus replication and introduced apical-out enteroids as a sensitive organoid-based system for studying sapovirus GI.1. Together, the introduced models provide complementary platforms that can contribute to knowledge about sapovirus pathogenesis and cell tropism as well as provide guidance in selecting suitable in vitro models for future sapovirus research.