Plant Disease Journal - 94(5):642 - Abstract
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Editor-in-Chief: Mark L. Gleason
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May 2010
ISSN: 0191-2917
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May 2010, Volume 94, Number 5
Page 642
http://dx.doi.org/10.1094/PDIS-94-5-0642C
Disease Notes
First Report of a Nanovirus Disease of Pea in
Germany
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I. Grigoras and B. Gronenborn, Institut des Sciences du Végétal, CNRS, 91198 Gif sur
Yvette, France; and H. J. Vetten, Julius Kühn Institute (JKI), Federal Research Center for
Cultivated Plants, Institute of Epidemiology and Pathogen Diagnostics, 38104
Braunschweig, Germany
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During the growing season of 2009, a disease consisting of leaf rolling, top yellows, and
plant stunting affected pea (Pisum sativum) in fields near Aschersleben, Saxony-Anhalt,
Germany. Samples from symptomatic plants collected in July 2009 were analyzed at the
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JKI in Braunschweig for infections by various legume viruses by ELISA, immunoelectron
microscopy, and transmission assays by sap and aphids. Of 23 samples, 9 were shown to
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contain Pea enation mosaic virus and three samples each contained Bean leafroll virus and
Soybean dwarf virus. From two further samples that had tested negative for the
aforementioned viruses, we succeeded in transferring a disease agent to faba bean (Vicia
faba) seedlings by giving 50 to 100 individuals of the pea aphid (Acyrthosiphon pisum)
acquisition and inoculation access feedings each of ~48 h. Following vector transmission,
the agent caused severe yellowing and stunting in pea and faba bean, sometimes followed
by necrosis. Attempts at mechanical transmission of the agent failed, and isolation of
double-stranded RNA from infected tissue was not successful. Therefore, we considered
the possible presence of a nanovirus (4). When using polyclonal antibodies (PAbs) against
Faba bean necrotic yellows virus (FBNYV) for double-antibody sandwich (DAS)-ELISA
analysis of the two isolates of the disease agent we observed weak but clearly positive
reactions. To confirm these weak DAS-ELISA reactions, we used all available monoclonal
antibodies (MAbs) raised against FBNYV (1) and faba bean necrotic stunt virus (FBNSV) (3)
individually in triple-antibody sandwich (TAS)-ELISA in combination with the FBNYV PAbs
for plate coating. Six of 26 MAbs reacted from weak to strong with the two pea isolates,
with MAbs FBNYV-3-1F7 and FBNSV-5-1G8 giving the strongest reactions and none of the
MAbs giving a differential reaction with the two pea isolates. Employing rolling circle
amplification of total DNA extracted from symptomatic leaves of one of the pea isolates
yielded a substantial amount of high molecular weight DNA, whereas little or no
amplification occurred when using DNA from noninoculated pea leaves. Restriction of the
amplified DNA in a nanovirus iteron-specific manner by AatII endonuclease yielded a
predominant and abundant product of ~1 kb (3). Sequence comparisons of eight cloned
DNAs of 1,002 nucleotides long unequivocally identified them as complete DNA-R
component of a new member of the genus Nanovirus (2,4). Its DNA-R sequence (GenBank
No. GU553134) is nearly equidistant from the DNA-R sequences of FBNYV (Y11405),
FBNSV (GQ150778), Milk vetch dwarf virus (MDV) (AB027511) and Subterranean clover
stunt virus (SCSV) (AJ290434), sharing with them respective sequence identities of 79,
78, 79, and 73%. Moreover, it is more distinct from the DNA-R sequences of FBNYV,
FBNSV, and MDV than the three latter are from each other (86 to 91%). This together with
the serological data relating to the capsid protein properties of this virus strongly suggest
that it is distinct from the hitherto described nanoviruses FBNYV, MDV, FBNSV, and SCSV.
Therefore, we propose the name pea necrotic yellow dwarf virus (PNYDV) for this new
nanovirus naturally infecting pea in Germany.
References: (1) A. Franz et al. Ann. Appl. Biol. 128:255, 1996. (2) I. Grigoras et al. J. Gen.
Virol. 89:583, 2008. (3) I. Grigoras et al. J. Virol. 83:10778, 2009. (4) H. J. Vetten et al.
Page 343 in: Virus Taxonomy. Elsevier/Academic Press, London, 2005.
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