Reverse genetics for crimean-congo hemorrhagic fever virus.
The widespread geographical distribution of Crimean-Congo hemorrhagic fever (CCHF) virus (more than 30 countries) and its ability to produce severe human disease with high mortality rates (up to 60%) make CCHF a major public health concern worldwide. We describe here the successful establishment of a reverse genetics technology for CCHF virus, a member of the genus Nairovirus, family BUNYAVIRIDAE: The RNA polymerase I (pol I) system was used to generate artificial viral RNA genome segments (minigenomes), which contained different reporter genes in antisense (virus RNA) or sense (virus-complementary RNA) orientation flanked by the noncoding regions of the CCHF virus S segment. Reporter gene expression was observed in different eukaryotic cell lines following transfection and subsequent superinfection with CCHF virus, confirming encapsidation, transcription, and replication of the pol I-derived minigenomes. The successful transfer of reporter gene activity to fresh cells demonstrated the generation of recombinant CCHF viruses, thereby confirming the packaging of the pol I-derived minigenomes into progeny viruses. The system offers a unique opportunity to study the biology of nairoviruses and to develop therapeutic and prophylactic measures against CCHF infections. In addition, we demonstrated for the first time that the human pol I system can be used to develop reverse genetics approaches for viruses in the family BUNYAVIRIDAE: This is important since it might facilitate the manipulation of bunyaviruses with cell and host tropisms restricted to primates.
Flick
Ramon
R
Special Pathogens Program, National Microbiology Laboratory, Health Canada, Canadian Science Centre for Human and Animal Health, Winnipeg, Canada R3E 3R2. Ramon_Flick@hc-sc.gc.ca
Flick
Kirsten
K
Feldmann
Heinz
H
Elgh
Fredrik
F
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D006224:Cricetinae; D005057:Eukaryotic Cells / Q000821:virology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D003225:Hemorrhagic Fever Virus, Crimean-Congo / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology; D006479:Hemorrhagic Fever, Crimean / Q000821:virology; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D010957:Plasmids; D012318:RNA Polymerase I / Q000378:metabolism*; D012367:RNA, Viral / Q000378:metabolism*; D011995:Recombination, Genetic; D014158:Transcription, Genetic; D014162:Transfection; D014779:Virus Replication
D008164:Luminescent Proteins; D012367:RNA, Viral; D049452:Green Fluorescent Proteins; D012318:RNA Polymerase I