Reverse genetics recovery of Lujo virus and role of virus RNA secondary structures in efficient virus growth.

Abstract:

:Arenaviruses are rodent-borne viruses with a bisegmented RNA genome. A genetically unique arenavirus, Lujo virus, was recently discovered as the causal agent of a nosocomial outbreak of acute febrile illness with hemorrhagic manifestations in Zambia and South Africa. The outbreak had a case fatality rate of 80%. A reverse genetics system to rescue infectious Lujo virus from cDNA was established to investigate the biological properties of this virus. Sequencing the genomic termini showed unique nucleotides at the 3' terminus of the S segment promoter element. While developing this system, we discovered that reconstructing infectious Lujo virus using the previously reported L segment intergenic region (IGR), comprising the arenaviral transcription termination signal, yielded an attenuated Lujo virus. Resequencing revealed that the correct L segment IGR was 36 nucleotides longer, and incorporating it into the reconstructed Lujo virus restored the growth rate to that of the authentic clinical virus isolate. These additional nucleotides were predicted to more than double the free energy of the IGR main stem-loop structure. In addition, incorporating the newly determined L-IGR into a replicon reporter system enhanced the expression of a luciferase reporter L segment. Overall, these results imply that an extremely stable secondary structure within the L-IGR is critical for Lujo virus propagation and viral protein production. The technology for producing recombinant Lujo virus now provides a method to precisely investigate the molecular determinants of virulence of this newly identified pathogen.

journal_name

J Virol

journal_title

Journal of virology

authors

Bergeron É,Chakrabarti AK,Bird BH,Dodd KA,McMullan LK,Spiropoulou CF,Nichol ST,Albariño CG

doi

10.1128/JVI.01144-12

subject

Has Abstract

pub_date

2012-10-01 00:00:00

pages

10759-65

issue

19

eissn

0022-538X

issn

1098-5514

pii

JVI.01144-12

journal_volume

86

pub_type

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