一种复制型单纯疱疹病毒 1（McKrae），其糖蛋白 K（gK）氨基末端的突变，使其在角膜感染后无法感染小鼠三叉神经节。

A replication competent HSV-1(McKrae) with a mutation in the amino-terminus of glycoprotein K (gK) is unable to infect mouse trigeminal ganglia after cornea infection.

机构信息

Division of Biotechnology and Molecular Medicine, Department of Pathobiological Sciences, School of Veterinary Medicine, Louisiana State University , Baton Rouge, LA , USA.

出版信息

Curr Eye Res. 2014 Jun;39(6):596-603. doi: 10.3109/02713683.2013.855238. Epub 2014 Jan 8.

DOI:10.3109/02713683.2013.855238

PMID:24401006

Abstract

PURPOSE

To determine the role of the amino terminus of herpes simplex virus-1 (HSV-1) glycoprotein K (gK) in corneal infection, neuroinvasion, and establishment of virus latency in trigeminal ganglia of mice.

METHODS

The recombinant virus HSV-1 (McKΔgK31-68) was constructed by engineering gK genes encoding gK lacking 38 amino acids immediately after the gK signal sequence. A rescued virus was also produced. Mouse eyes were scarified and infected with 10(5) plaque forming units (PFU) in each eye. Clinical signs of ocular disease were monitored daily. Thirty days postinfection trigeminal ganglia were collected and processed for quantitative PCR (qPCR) analysis of viral DNA and recovery of infectious virions by cell culture of ganglionic tissues.

RESULTS

Deletion of the amino terminus of gK encoded by the McKΔgK31-68 mutant virus did not substantially affect its replication kinetics on African green monkey kidney cells (Vero), while it reduced cell-to-cell spread. McK viral infection of scarified mouse corneas with 10(5) PFU produced severe ocular disease. In contrast, McKΔgK31-68 viral infection with 10(5) PFU produced no significant ocular disease symptoms. All ganglia from mice infected with the McK virus produced high numbers of infectious virions upon explant culture in Vero cells, in agreement with qPCR results detecting high number of HSV-1 viral DNA in ganglionic tissues. In contrast, qPCR failed to detect any viral genomes in McKΔgK31-68 ganglia, while two of the ten ganglia revealed the presence of low numbers of infectious virions upon explant culture in Vero cells.

CONCLUSIONS

The results show that the amino terminus of gK is essential for neuroinvasiveness and acute herpes keratitis in the mouse eye model. It is likely that gK is involved in efficient infection of axonal termini, since mouse eye scarification provided a direct access to the high density of neuronal axons innervating mouse corneas.

摘要

目的

确定单纯疱疹病毒 1 型（HSV-1）糖蛋白 K（gK）氨基末端在小鼠三叉神经节中角膜感染、神经侵袭和病毒潜伏建立中的作用。

方法

通过工程化 gK 基因构建重组病毒 HSV-1（McKΔgK31-68），该基因编码 gK 信号序列后立即缺失 38 个氨基酸。还产生了一种拯救病毒。用 10（5）噬菌斑形成单位（PFU）在每只眼睛中划伤和感染小鼠眼睛。每天监测眼部疾病的临床症状。感染后 30 天收集三叉神经节并进行定量 PCR（qPCR）分析病毒 DNA 和通过神经节组织的细胞培养回收感染性病毒粒子。

结果

McKΔgK31-68 突变病毒氨基末端缺失对其在非洲绿猴肾细胞（Vero）上的复制动力学没有显著影响，但降低了细胞间传播。用 10（5）PFU 划伤的小鼠角膜感染 McK 病毒可引起严重的眼部疾病。相比之下，用 10（5）PFU 感染 McKΔgK31-68 病毒不会引起明显的眼部疾病症状。在用 McK 病毒感染的所有小鼠神经节中，在用 Vero 细胞进行外植体培养时产生大量感染性病毒粒子，这与 qPCR 检测到神经节组织中高数量 HSV-1 病毒 DNA 的结果一致。相比之下，qPCR 未能在 McKΔgK31-68 神经节中检测到任何病毒基因组，而在 10 个神经节中的两个中，在用 Vero 细胞进行外植体培养时检测到低数量的感染性病毒粒子。