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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

Defective influenza A virus generated entirely from plasmids: its RNA is expressed in infected mouse lung and modulates disease.

Naturally produced defective influenza virus has antiviral activity and, in sufficient amount, can protect mice from lethal influenza, irrespective of the virus subtype causing the disease. However, such defective virus preparations contain many undefined defective RNA sequences, and it is thus not possible to establish dose-response relationships. To address this situation, we have transfected DNA encoding a cloned defective RNA into Vero cells along with the 17 A/WSN (H1N1) plasmids required for infectious helper virus, and produced molecularly cloned defective virus. Here we used POLI-220 that expresses a 445 nt defective RNA isolated from a mouse-protective defective equine H3N8 virus, and POLI-317 that expresses a 585 nt defective RNA from an avian H7N7 virus. Both originate from genomic segment 1. Virus preparations were UV-irradiated selectively to destroy virus infectivity but not the activity of the defective RNAs, and adult mice were inoculated intranasally with defective virus and WSN (H1N1) challenge virus (10 LD(50)). Defective POLI-220 and POLI-317 RNAs were detected readily in infected lung tissue by RT-PCR, but these Vero cell preparations did not modulate disease. However, after a single passage in embryonated eggs, defective POLI-220 and POLI-317 viruses significantly delayed the onset of disease and death in WSN-infected mice, although did not affect final mortality. Direct PCR sequencing confirmed the identity of mouse-passaged defective RNAs and showed that none had undergone any sequence changes. With this advance it will now be possible to study the interference phenomenon in vivo with defective viruses carrying a defined defective RNA.[1]


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