Human noroviruses (HuNoVs) are the leading cause of epidemic and sporadic acute gastroenteritis worldwide. We previously demonstrated human intestinal stem cell-derived enteroids (HIEs) support cultivation of several HuNoV strains. However, HIEs did not support virus replication from every HuNoV-positive stool sample, which led us to test and optimize new medium conditions, identify characteristics of stool samples that allow replication, and evaluate consistency of replication over time. Optimization of our HIE-HuNoV culture system has shown the following: (i) a new HIE culture medium made with conditioned medium from a single cell line and commercial media promotes robust replication of HuNoV strains that replicated poorly in HIEs grown in our original culture medium made with conditioned media from 3 separate cell lines; (ii) GI.1, 11 GII genotypes (GII.1, GII.2, GII.3, GII.4, GII.6, GII.7, GII.8, GII.12, GII.13, GII.14, and GII.17), and six GII.4 variants can be cultivated in HIEs; (iii) successful replication is more likely with virus in stools with higher virus titers; (iv) GII.4_Sydney_2012 virus replication was reproducible over 3 years; and (v) HuNoV infection is restricted to the small intestine, based on replication of two viral strains in duodenal and ileal HIEs, but not colonoids, from two susceptible donors. These results improve the HIE culture system for HuNoV replication. Use of HIEs by several laboratories worldwide to study the molecular mechanisms that regulate HuNoV replication confirms the usefulness of this culture system, and our optimized methods for virus replication will advance the development of effective therapies and methods for virus control.

中文翻译：

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人类肠道小肠中的新见解和增强人类诺如病毒培养

人类诺如病毒 (HuNoVs) 是全球流行性和散发性急性胃肠炎的主要原因。我们之前证明了人类肠道干细胞衍生的小肠 (HIE) 支持几种 HuNoV 菌株的培养。然而，HIE 不支持从每个 HuNoV 阳性粪便样本中复制病毒，这导致我们测试和优化新的培养基条件，确定允许复制的粪便样本的特征，并评估复制随时间的一致性。我们的 HIE-HuNoV 培养系统的优化显示如下：(i) 用来自单细胞系的条件培养基和商业培养基制成的新 HIE 培养基促进了在我们原始培养基中生长的 HIE 中复制较差的 HuNoV 菌株的稳健复制用来自 3 个独立细胞系的条件培养基制成；(ii) GI.1，11 种 GII 基因型（GII.1、GII.2、GII.3、GII.4、GII.6、GII.7、GII.8、GII.12、GII.13、GII.14 和 GII.17），并且可以在 HIE 中培养 6 个 GII.4 变体；(iii) 粪便中病毒滴度较高的病毒更可能成功复制；(iv) GII.4_Sydney_2012 病毒复制可在 3 年内重现；(v) HuNoV 感染仅限于小肠，基于来自两个易感供体的两种病毒株在十二指肠和回肠 HIE 中的复制，但不是在结肠样中。这些结果改进了用于 HuNoV 复制的 HIE 培养系统。全球多个实验室使用 HIE 研究调节 HuNoV 复制的分子机制证实了这种培养系统的有用性，我们优化的病毒复制方法将推动开发有效的病毒控制疗法和方法。