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Journal of Virology, October 2009, p. 10778-10787, Vol. 83, No. 20
0022-538X/09/$08.00+0     doi:10.1128/JVI.01212-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Reconstitution of Authentic Nanovirus from Multiple Cloned DNAs {triangledown} ,{dagger}

Ioana Grigoras,1 Tatiana Timchenko,1 Lina Katul,2 Ana Grande-Pérez,3 Heinrich-Josef Vetten,2 and Bruno Gronenborn1*

Institut des Sciences du Végétal, CNRS, 91198 Gif sur Yvette, France,1 Julius Kühn Institute, Bundesforschungsinstitut für Kulturpflanzen, Institut für Epidemiologie und Pathogendiagnostik, 38104 Braunschweig, Germany,2 Universidad de Málaga, Area de Genética, 29071 Málaga, Spain3

Received 12 June 2009/ Accepted 30 July 2009

We describe a new plant single-stranded DNA (ssDNA) virus, a nanovirus isolate originating from the faba bean in Ethiopia. We applied rolling circle amplification (RCA) to extensively copy the individual circular DNAs of the nanovirus genome. By sequence analyses of more than 208 individually cloned genome components, we obtained a representative sample of eight polymorphic swarms of circular DNAs, each about 1 kb in size. From these heterogeneous DNA populations after RCA, we inferred consensus sequences of the eight DNA components of the virus genome. Based on the distinctive molecular and biological properties of the virus, we propose to consider it a new species of the genus Nanovirus and to name it faba bean necrotic stunt virus (FBNSV). Selecting a representative clone of each of the eight DNAs for transfer by T-DNA plasmids of Agrobacterium tumefaciens into Vicia faba plants, we elicited the development of the typical FBNSV disease symptoms. Moreover, we showed that the virus thus produced was readily transmitted by two different aphid vector species, Aphis craccivora and Acyrthosiphon pisum. This represents the first reconstitution of a fully infectious and sustainably insect-transmissible nanovirus from its cloned DNAs and provides compelling evidence that the genome of a legume-infecting nanovirus is typically comprised of eight distinct DNA components.

* Corresponding author. Mailing address: Institut des Sciences du Végétal, CNRS, Avenue de la Terrasse, Bât. 23, 91198 Gif sur Yvette, France. Phone: 33 169823833. Fax: 33 169823695. E-mail: gronenborn{at}isv.cnrs-gif.fr

{triangledown} Published ahead of print on 5 August 2009.

{dagger} Supplemental material for this article may be found at http://jvi.asm.org/.

Journal of Virology, October 2009, p. 10778-10787, Vol. 83, No. 20
0022-538X/09/$08.00+0     doi:10.1128/JVI.01212-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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