Emergence of a Large-Plaque Variant in Mice Infected with Coxsackievirus B3.
Emergence of a Large-Plaque Variant in Mice Infected with Coxsackievirus B3.
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DOI：
10.1128/mbio.00119-16
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发表时间：
2016-03-29
期刊：
影响因子：
6.4
通讯作者：
Pfeiffer JK
中科院分区：
文献类型：
--
作者：
Wang Y;Pfeiffer JK
柯萨奇病毒是经常感染人类的肠道病毒。为了研究柯萨奇病毒的发病机制，我们口服接种小鼠与柯萨奇病毒B3（CVB 3）南希株。使用琼脂覆盖的HeLa细胞空斑测定，我们注意到一些粪便病毒产生的空斑大于接种病毒的100倍。这些大空斑变异体在几种不同组织中病毒复制后出现。我们确定了一个单一的氨基酸变化，N63 Y，在VP 3衣壳蛋白，这是足以赋予大空斑表型。当琼脂糖代替琼脂用于覆盖时，野生型CVB 3和N63 Y突变型CVB 3具有相似的噬斑大小。我们确定琼脂中的硫酸化聚糖抑制野生型CVB 3的噬斑形成，但不抑制N63 Y突变型CVB 3的噬斑形成。此外，N63 Y突变CVB 3结合肝素（一种硫酸化聚糖）的效率低于野生型CVB 3。虽然N63 Y突变型CVB 3在培养细胞中具有生长缺陷和附着减少，但它在小鼠中具有增强的复制和致病性。感染N63 Y突变型CVB 3比感染野生型CVB 3诱导更严重的肝损伤，可能是因为N63 Y突变型CVB 3更有效地传播到肝脏。我们的数据强化了这样一种观点，即培养适应的实验室病毒株在体内的适应性可能会降低。N63 Y突变CVB 3可能是一个有用的平台，以了解病毒的适应和发病机制，在动物研究。柯萨奇病毒经常感染人类，尽管许多感染是轻度或无症状的，但可能会有严重的后果，包括心脏炎症。大多数关于柯萨奇病毒和其他病毒的研究使用实验室适应的病毒株，因为它们在细胞培养中有效复制。我们使用细胞培养适应株CVB 3，南希，检查病毒复制和发病机制，在口服接种小鼠。我们发现，小鼠的病毒脱落与输入病毒不同，因为它们在细胞培养中形成了非常大的噬斑。我们发现了一个单一的突变，VP 3 N63 Y，这是足够的大斑块形成。N63 Y突变病毒在细胞培养物中的聚糖结合和复制减少;然而，它们在小鼠中的复制和毒力增强。我们现在使用N63 Y突变CVB 3作为病毒致病机制研究的改进系统。
Coxsackieviruses are enteric viruses that frequently infect humans. To examine coxsackievirus pathogenesis, we orally inoculated mice with the coxsackievirus B3 (CVB3) Nancy strain. Using HeLa cell plaque assays with agar overlays, we noticed that some fecal viruses generated plaques >100 times as large as inoculum viruses. These large-plaque variants emerged following viral replication in several different tissues. We identified a single amino acid change, N63Y, in the VP3 capsid protein that was sufficient to confer the large-plaque phenotype. Wild-type CVB3 and N63Y mutant CVB3 had similar plaque sizes when agarose was used in the overlay instead of agar. We determined that sulfated glycans in agar inhibited plaque formation by wild-type CVB3 but not by N63Y mutant CVB3. Furthermore, N63Y mutant CVB3 bound heparin, a sulfated glycan, less efficiently than wild-type CVB3 did. While N63Y mutant CVB3 had a growth defect in cultured cells and reduced attachment, it had enhanced replication and pathogenesis in mice. Infection with N63Y mutant CVB3 induced more severe hepatic damage than infection with wild-type CVB3, likely because N63Y mutant CVB3 disseminates more efficiently to the liver. Our data reinforce the idea that culture-adapted laboratory virus strains can have reduced fitness in vivo. N63Y mutant CVB3 may be useful as a platform to understand viral adaptation and pathogenesis in animal studies. Coxsackieviruses frequently infect humans, and although many infections are mild or asymptomatic, there can be severe outcomes, including heart inflammation. Most studies with coxsackieviruses and other viruses use laboratory-adapted viral strains because of their efficient replication in cell culture. We used a cell culture-adapted strain of CVB3, Nancy, to examine viral replication and pathogenesis in orally inoculated mice. We found that mice shed viruses distinct from input viruses because they formed extremely large plaques in cell culture. We identified a single mutation, VP3 N63Y, that was sufficient for large-plaque formation. N63Y mutant viruses have reduced glycan binding and replication in cell culture; however, they have enhanced replication and virulence in mice. We are now using N63Y mutant CVB3 as an improved system for viral pathogenesis studies.