Oka Tomoichiro, Stoltzfus Garrett T, Zhu Chelsea, Jung Kwonil, Wang Qiuhong, Saif Linda J
Food Animal Health Research Program, Ohio Agricultural Research and Development Center, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, OH, United States of America.
Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
PLoS One. 2018 Feb 13;13(2):e0178157. doi: 10.1371/journal.pone.0178157. eCollection 2018.
Noroviruses (NoVs) and Sapoviruses (SaVs) are enteric caliciviruses that have been detected in multiple mammalian species, including humans. Currently, efficient cell culture systems have been established only for murine NoVs and porcine SaV Cowden strain. Establishment of an efficient in vitro cell culture system for other NoVs and SaVs remains challenging; however, human NoV (HuNoV) replication in 3D cultured Caco-2 cells and a clone of Caco-2 cells, C2BBe1, human enteroids and in human B cells has been reported. In this study, we tested various cells and culture conditions to grow HuNoVs and a human SaV (HuSaV) to test the possibility of the propagation in different cells and culture conditions. We also attempted to grow a bovine NoV (BoNoV) in ex vivo organ cultures. We did not observe significant RNA level increases for HuSaV and BoNoV under our test conditions. HuNoV RNA levels increased to a maximum of ~600-fold in long-term Caco-2 cells that were cultured for 1-2 months in multi-well plates and inoculated with HuNoV-positive and bacteria-free human stool suspensions using serum-free medium supplemented with the bile acid, GCDCA. However, this positive result was inconsistent. Our results demonstrated that HuNoVs, BoNoV and HuSaV largely failed to grow in vitro under our test conditions. Our purpose is to share our findings with other researchers with the goal to develop efficient, reproducible simplified and cost-effective culture systems for human and animal NoVs and SaVs in the future.
诺如病毒（NoVs）和札如病毒（SaVs）是肠道杯状病毒，已在包括人类在内的多种哺乳动物物种中被检测到。目前，仅针对鼠诺如病毒和猪札如病毒考登株建立了有效的细胞培养系统。建立针对其他诺如病毒和札如病毒的高效体外细胞培养系统仍然具有挑战性；然而，已有报道称人诺如病毒（HuNoV）可在三维培养的Caco-2细胞、Caco-2细胞系的一个克隆C2BBe1、人肠道类器官以及人B细胞中复制。在本研究中，我们测试了多种细胞和培养条件以培养HuNoV和一种人札如病毒（HuSaV），以检验在不同细胞和培养条件下进行增殖的可能性。我们还尝试在离体器官培养物中培养牛诺如病毒（BoNoV）。在我们的测试条件下，我们未观察到HuSaV和BoNoV的RNA水平有显著升高。在多孔板中培养1至2个月的长期Caco-2细胞中，使用补充了胆汁酸甘氨胆酸（GCDCA）的无血清培养基接种HuNoV阳性且无菌的人类粪便悬液后，HuNoV RNA水平最高增加至约600倍。然而，这一阳性结果并不一致。我们的结果表明，在我们的测试条件下，HuNoV、BoNoV和HuSaV在体外大多无法生长。我们的目的是与其他研究人员分享我们的发现，以期未来开发出高效、可重复、简化且具有成本效益的用于人和动物诺如病毒及札如病毒的培养系统。
