Flaviviruses are representative arboviruses carried by arthropods and/or vertebrates; these viruses can pose a public health concern in many countries. By contrast, it is known that a novel virus group called insect-specific flaviviruses (ISFs) also infects arthropods, although no such virus has yet been isolated from vertebrates. The characteristics of ISFs, which affect replication of human-pathogenic flaviviruses within co-infected mosquito cells or mosquitoes without affecting the mosquitoes themselves, mean that we should pay attention to both ISFs and human-pathogenic flaviviruses, despite the fact that ISFs appear not to be directly hazardous to human health. To assess the risk of diseases caused by flaviviruses, and to better understand their ecology, it is necessary to know the extent to which flaviviruses are harbored by arthropods. We developed a novel universal primer for use in a PCR-based system to detect a broad range of flaviviruses. We then evaluated its performance. The utility of the novel primer pair was evaluated in a PCR assay using artificially synthesized oligonucleotides derived from a template viral genome sequence. The utility of the primer pair was also examined by reverse transcription PCR (RT-PCR) using cDNA templates prepared from virus-infected cells or crude supernatants prepared from virus-containing mosquito homogenates. The novel primer pair amplified the flavivirus NS5 sequence (artificially synthesized) in all samples tested (six species of flavivirus that can cause infectious diseases in humans, and flaviviruses harbored by insects). In addition, the novel primer pair detected viral genomes in cDNA templates prepared from mosquito cells infected with live flavivirus under different infectious conditions. Finally, the viral genome was detected with high sensitivity in crude supernatants prepared from pooled mosquito homogenates. This PCR system based on a novel primer pair makes it possible to detect arthropod-borne flaviviruses worldwide (the primer pair even detected viruses belonging to different genetic subgroups). As such, an assay based on this primer pair may help to improve public health and safety, as well as increase our understanding of flavivirus ecology.

中文翻译：

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开发用于检测黄病毒的属特异性通用引物

黄病毒是节肢动物和/或脊椎动物携带的代表性虫媒病毒；这些病毒可能在许多国家造成公共卫生问题。相比之下，已知一种称为昆虫特异性黄病毒 (ISF) 的新型病毒群也会感染节肢动物，尽管尚未从脊椎动物中分离出此类病毒。ISFs 的特性会影响人类致病性黄病毒在共同感染的蚊子细胞或蚊子中的复制而不影响蚊子本身，这意味着我们应该同时关注 ISFs 和人类致病性黄病毒，尽管事实上 ISFs 似乎没有直接危害人体健康。为了评估由黄病毒引起的疾病的风险，并更好地了解它们的生态学，有必要了解节肢动物携带黄病毒的程度。我们开发了一种新型通用引物，可用于基于 PCR 的系统，以检测范围广泛的黄病毒。然后我们评估了它的性能。使用源自模板病毒基因组序列的人工合成寡核苷酸，在 PCR 分析中评估了新型引物对的效用。还使用从病毒感染的细胞制备的 cDNA 模板或从含病毒的蚊子匀浆制备的粗上清液通过逆转录 PCR (RT-PCR) 检查了引物对的效用。新的引物对在所有测试样品（可引起人类传染病的六种黄病毒和昆虫携带的黄病毒）中扩增了黄病毒 NS5 序列（人工合成）。此外，新的引物对在不同感染条件下用活黄病毒感染的蚊子细胞制备的 cDNA 模板中检测到病毒基因组。最后，在由混合的蚊子匀浆制备的粗上清液中以高灵敏度检测病毒基因组。这种基于新型引物对的 PCR 系统可以在全球范围内检测节肢动物传播的黄病毒（引物对甚至检测到属于不同遗传亚群的病毒）。因此，基于该引物对的分析可能有助于改善公众健康和安全，并增加我们对黄病毒生态学的了解。这种基于新型引物对的 PCR 系统可以在全球范围内检测节肢动物传播的黄病毒（引物对甚至检测到属于不同遗传亚群的病毒）。因此，基于该引物对的分析可能有助于改善公众健康和安全，并增加我们对黄病毒生态学的了解。这种基于新型引物对的 PCR 系统可以在全球范围内检测节肢动物传播的黄病毒（引物对甚至检测到属于不同遗传亚群的病毒）。因此，基于该引物对的分析可能有助于改善公众健康和安全，并增加我们对黄病毒生态学的了解。