Simón Oihane, Williams Trevor, López-Ferber Miguel, Caballero Primitivo
Departamento de Producción Agraria, Universidad Pública de Navarra, 31006 Pamplona, Spain.
Appl Environ Microbiol. 2004 Sep;70(9):5579-88. doi: 10.1128/AEM.70.9.5579-5588.2004.
A Nicaraguan field isolate (SfNIC) of Spodoptera frugiperda nucleopolyhedrovirus was purified by plaque assay on Sf9 cells. Nine distinct genotypes, A to I, were identified by their restriction endonuclease profiles. Variant SfNIC-B was selected as the standard because its restriction profile corresponded to that of the wild-type isolate. Physical maps were generated for each of the variants. The differences between variants and the SfNIC-B standard were confined to the region between map units 9 and 32.5. This region included PstI-G, PstI-F, PstI-L, PstI-K and EcoRI-L fragments. Eight genotypes presented a deletion in their genome compared with SfNIC-B. Occlusion body-derived virions of SfNIC-C, -D and -G accounted for 41% of plaque-purified clones. These variants were not infectious per os but retained infectivity by injection into S. frugiperda larvae. Median 50% lethal concentration values for the other cloned genotypes were significantly higher than that of the wild type. The variants also differed in their speed of kill. Noninfectious variants SfNIC-C and -D lacked the pif and pif-2 genes. Infectivity was restored to these variants by plasmid rescue with a plasmid comprising both pif and pif-2. Transcription of an SfNIC-G gene was detected by reverse transcription-PCR in insects, but no fatal disease developed. Transcription was not detected in SfNIC-C or -D-inoculated larvae. We conclude that the SfNIC population presents high levels of genetic diversity, localized to a 17-kb region containing pif and pif-2, and that interactions among complete and deleted genotypic variants will likely influence the capacity of this virus to control insect pests.
通过在 Sf9 细胞上进行蚀斑测定，纯化了草地贪夜蛾核多角体病毒的尼加拉瓜田间分离株（SfNIC）。根据其限制性内切酶图谱鉴定出了 9 种不同的基因型，A 至 I。选择变体 SfNIC-B 作为标准，因为其限制性图谱与野生型分离株的图谱一致。为每个变体生成了物理图谱。变体与 SfNIC-B 标准之间的差异局限于图谱单位 9 和 32.5 之间的区域。该区域包括 PstI-G、PstI-F、PstI-L、PstI-K 和 EcoRI-L 片段。与 SfNIC-B 相比，8 种基因型在其基因组中存在缺失。SfNIC-C、-D 和 -G 的包涵体衍生病毒粒子占蚀斑纯化克隆的 41%。这些变体经口不具有感染性，但通过注射到草地贪夜蛾幼虫中仍保留感染性。其他克隆基因型的半数致死浓度中值显著高于野生型。这些变体在致死速度上也有所不同。无感染性的变体 SfNIC-C 和 -D 缺乏 pif 和 pif-2 基因。用包含 pif 和 pif-2 的质粒进行质粒拯救可恢复这些变体的感染性。通过逆转录 PCR 在昆虫中检测到了 SfNIC-G 基因的转录，但未引发致命疾病。在接种 SfNIC-C 或 -D 的幼虫中未检测到转录。我们得出结论，SfNIC 群体呈现出高水平的遗传多样性，定位于包含 pif 和 pif-2 的 17 kb 区域，并且完整和缺失基因型变体之间的相互作用可能会影响该病毒控制害虫的能力。
