Flipsen J T, Martens J W, van Oers M M, Vlak J M, van Lent J W
Department of Virology, Wageningen Agricultural University, The Netherlands.
Virology. 1995 Apr 1;208(1):328-35. doi: 10.1006/viro.1995.1156.
A special recombinant of Autographa californica multicapsid nuclear polyhedrosis virus (AcNPV) was designed to study the early histopathological events of baculovirus infection in Spodoptera exigua larvae. This recombinant contained a Drosophila melanogaster heat shock 70 promoter driving an Escherichia coli beta-galactosidase (Lac-Z) reporter gene to monitor the presence of early viral gene expression and a second reporter gene, the E. coli beta-glucuronidase (GUS) gene, under control of the very late AcNPV p10 promoter to monitor viral replication. In S. exigua larvae, permissive Spodoptera spp. cultured cells, and nonpermissive D. melanogaster cultured cells early viral gene expression was indicated by the appearance of Lac-Z as early as 3 hr p.i. Late viral gene expression was indicated by the appearance of GUS and occurred only in the permissive cultured cells and larvae. Early and late viral gene expression could be detected simultaneously using differential enzyme histochemistry. Analysis of infected S. exigua larvae revealed that midgut columnar cells and, at a low frequency, midgut regenerative cells were the primary sites of infection. Parental nucleocapsids were apparently transported through columnar cells to underlaying regenerative cells before virus replication and progeny production. Infection of tissues beside the midgut epithelium was not detected prior to viral replication within the midgut, suggesting that infection of the midgut is an important prelude to systemic infection.
设计一种苜蓿银纹夜蛾多粒包埋核型多角体病毒（AcNPV）的特殊重组体，以研究杆状病毒感染甜菜夜蛾幼虫的早期组织病理学事件。该重组体包含一个驱动大肠杆菌β-半乳糖苷酶（Lac-Z）报告基因的黑腹果蝇热休克70启动子，用于监测早期病毒基因表达的存在，以及第二个报告基因，即大肠杆菌β-葡萄糖醛酸酶（GUS）基因，受极晚期AcNPV p10启动子控制，用于监测病毒复制。在甜菜夜蛾幼虫、允许的夜蛾属培养细胞和不允许的黑腹果蝇培养细胞中，早在感染后3小时Lac-Z的出现就表明了早期病毒基因表达。GUS的出现表明了晚期病毒基因表达，且仅发生在允许的培养细胞和幼虫中。使用差异酶组织化学可以同时检测早期和晚期病毒基因表达。对受感染的甜菜夜蛾幼虫的分析表明，中肠柱状细胞以及低频出现的中肠再生细胞是主要感染部位。亲代核衣壳显然在病毒复制和子代产生之前通过柱状细胞转运到下层的再生细胞。在中肠内病毒复制之前未检测到中肠上皮以外组织的感染，这表明中肠感染是全身感染的重要前奏。
