Astrovirus VA1/HMO-C (VA1; mamastrovirus 9) is a recently discovered astrovirus genotype that is divergent from the classic human astroviruses (mamastrovirus 1). The gastrointestinal tract is presumed to be the primary site of infection and pathogenicity for astroviruses. However, VA1 has been independently detected in brain tissue of five cases of human encephalitis. Studies of the pathogenicity of VA1 are currently impossible because there are no reported cell culture systems or in vivo models that support VA1 infection. Here, we describe successful propagation of VA1 in multiple human cell lines. The initial inoculum, a filtered clinical stool sample from the index gastroenteritis case cluster that led to the discovery of VA1, was first passaged in Vero cells. Serial blind passage in Caco-2 cells yielded increasing copies of VA1 RNA, and multistep growth curves demonstrated a >100-fold increase in VA1 RNA 72 h after inoculation. The full-length genomic and subgenomic RNA strands were detected by Northern blotting, and crystalline lattices of viral particles of ∼26-nm diameter were observed by electron microscopy in infected Caco-2 cells. Unlike other human astrovirus cell culture systems, which require addition of exogenous trypsin for continued propagation, VA1 could be propagated equally well with or without the addition of trypsin. Furthermore, VA1 was sensitive to the type I interferon (IFN-I) response, as VA1 RNA levels were reduced by pretreatment of Caco-2 cells with IFN-β1a. The ability to propagate VA1 in cell culture will facilitate studies of the neurotropism and neuropathogenesis of VA1.

IMPORTANCE Astroviruses are an emerging cause of central nervous system infections in mammals, and astrovirus VA1/HMO-C is the most prevalent astrovirus in cases of human encephalitis. This virus has not been previously propagated, preventing elucidation of the biology of this virus. We describe the first cell culture system for VA1, a key step necessary for the study of its ability to cause disease.

星状病毒VA1 / HMO-C（VA1；乳腺病毒9）是最近发现的一种星型病毒基因型，它与经典的人类星状病毒（乳腺病毒1）不同。胃肠道被认为是星形病毒感染和致病性的主要部位。但是，已经在五例人类脑炎的脑组织中独立检测到VA1。目前尚无关于VA1致病性的研究，因为目前尚无细胞培养系统或体内报道支持VA1感染的模型。在这里，我们描述了VA1在多个人类细胞系中的成功繁殖。最初的接种物首先在Vero细胞中传代，这是从索引性肠胃炎病例簇中过滤出来的临床粪便样本，导致发现了VA1。在Caco-2细胞中连续盲传产生了VA1 RNA拷贝的增加，并且多步生长曲线表明接种72 h后VA1 RNA的> 100倍增加。通过Northern印迹检测全长基因组和亚基因组RNA链，并通过电子显微镜在感染的Caco-2细胞中观察到直径约26nm的病毒颗粒的晶格。与其他人类星状病毒细胞培养系统不同，后者需要添加外源胰蛋白酶才能继续繁殖，在添加或不添加胰蛋白酶的情况下，VA1均可以很好地繁殖。此外，VA1对I型干扰素（IFN-I）反应敏感，因为通过用IFN-β1a预处理Caco-2细胞可降低VA1 RNA水平。在细胞培养物中繁殖VA1的能力将促进VA1的神经向性和神经发病机制的研究。

重要性星状病毒在哺乳动物中枢神经系统感染的一个新兴的原因，星状病毒VA1 / HMO-C是人类脑炎病例中最常见的星状病毒。该病毒以前未曾传播过，从而无法阐明该病毒的生物学特性。我们描述了第一个VA1细胞培养系统，这是研究其致病能力所必需的关键步骤。