Center for Food Safety, Department of Food Science and Technology, University of Georgia, Griffin, GA 30223, USA.
Viruses. 2024 Feb 2;16(2):241. doi: 10.3390/v16020241.
Human intestinal enteroids (HIEs) culture is an emerging model for assessing the infectivity of human noroviruses (HuNoVs). The model is based on detecting an increase in HuNoV RNA post-infection of HIEs. HuNoV fecal samples used for HIE infection are traditionally processed by serial filtration. Recently, processing HuNoV fecal samples by serial centrifugation was shown to retain vesicles containing HuNoV. The objective of this study was to investigate whether serially centrifuged fecal samples, RNA extraction kit (QIAamp versus MagMaX) and HIE age (newer versus older) affect HuNoV RNA fold increase in HIE. HuNoV GII.1, GII.4 and GII.6 fecal samples were prepared by serial centrifugation and filtration and the viral RNA in HIE was quantified at 1 and 72 h post-infection (hpi) following RNA extraction and RT-qPCR. The serially filtered GII.1, GII.4 and GII.6 showed successful replication in HIE, resulting in mean log increases of 2.2, 2 and 1.2, respectively, at 72 vs. 1 hpi. In contrast, only serially centrifuged GII.1 showed consistently successful replication. However, using newer HIE passages and the MagMAX kit resulted in mean log fold increases for serially centrifuged GII.1, GII.4 and GII.6 (1.6, 2.3 and 1.8 log, respectively) that were similar to serially filtered samples. Therefore, HuNoV fecal sample processing and HIE age can affect virus replication in the HIE model.
人类肠道类器官（HIEs）培养是评估人类诺如病毒（HuNoVs）感染性的新兴模型。该模型基于感染 HIEs 后 HuNoV RNA 的增加来检测。用于 HIE 感染的 HuNoV 粪便样本传统上通过连续过滤进行处理。最近，研究表明，通过连续离心处理 HuNoV 粪便样本可以保留含有 HuNoV 的囊泡。本研究旨在研究连续离心的粪便样本、RNA 提取试剂盒（QIAAmp 与 MagMaX）和 HIE 年龄（较新与较旧）是否会影响 HIE 中 HuNoV RNA 的倍数增加。通过连续离心和过滤制备 HuNoV GII.1、GII.4 和 GII.6 粪便样本，并在 RNA 提取和 RT-qPCR 后，于感染后 1 和 72 小时（hpi）定量 HIE 中的病毒 RNA。连续过滤的 GII.1、GII.4 和 GII.6 在 HIE 中成功复制，导致 72 hpi 与 1 hpi 相比，平均对数增加分别为 2.2、2 和 1.2。相比之下，只有连续离心的 GII.1 显示出一致的成功复制。然而，使用较新的 HIE 传代和 MagMAX 试剂盒，连续离心的 GII.1、GII.4 和 GII.6 的平均对数倍数增加（分别为 1.6、2.3 和 1.8 对数）与连续过滤的样本相似。因此，HuNoV 粪便样本处理和 HIE 年龄可能会影响 HIE 模型中的病毒复制。
