Co-Infection of Tobacco Rattle and Cycas Necrotic Stunt Viruses in Paeonia lactiflora: Detection Strategies, Potential Origins of Infection, and Implications for Paeonia Germplasm Conservation.

Increasing reports of tobacco rattle virus (TRV) and cycas necrotic stunt virus (CNSV) in herbaceous Paeonia worldwide highlight the importance of conserving the genetic resources of this economically important ornamental and medicinal crop. The unknown origin(s) of infection, differential susceptibility of peony cultivars to these viruses, and elusive disease phenotypes for CNSV in peonies make early detection and management challenging. Here, we report the presence of TRV and CNSV in plants of the University of Michigan living peony collection in the United States and a molecular characterization of their strains. Using sequences of the TRV 194 K RNA polymerase gene, we confirmed TRV infections in seven symptomatic plants (1.07% of all plants in the collection). Using newly developed primers, we recovered sequences of the CNSV RdRp gene and the polyprotein 1 gene region from nine out of twelve samples analyzed, including three from symptomless plants. Four of the nine plants had TRV and CNSV co-infections and showed more severe disease symptoms than plants only infected with TRV. Phylogenetic analyses of isolates from the University of Michigan living peony collection and publicly available isolates point to multiple origins of TRV and CNSV infections in this collection. This is the first report of TRV/CNSV co-infection and of a symptomatic detection of CNSV on cultivated P. lactiflora.

Complete genome sequence of peony leafroll-associated virus, a novel ampelovirus in subgroup I.

The complete genome sequence of peony leafroll-associated virus (PLRaV) was determined by deep sequencing of ribosomal-RNA-depleted total RNA extracted from a peony plant exhibiting leafroll symptoms. Further PCR and RACE analysis showed that the PLRaV genome consists of 15,406 nucleotides and contains 10 putative open reading frames, with an organization typical of members of the genus Ampelovirus, family Closteroviridae. Amino acid sequence comparisons showed that the viral heat shock protein 70 homolog (HSP70h) shared the highest sequence identity (41.7%) with the corresponding region of grapevine leafroll-associated virus 1, and the coat protein (CP) and RNA-dependent RNA polymerase (RdRp) shared the highest sequence identity (32.1% and 52.3%, respectively) with grapevine leafroll-associated virus 13. Phylogenetic analysis of the HSP70h, CP, and RdRp aa sequences showed that PLRaV clustered with members of subgroup I of the genus Ampelovirus.

Genomic detection and molecular characterization of two distinct isolates of cycas necrotic stunt virus from Paeonia suffruticosa and Daphne odora.

Complete genome sequences of two cycas necrotic stunt virus (CNSV) isolates from Paeonia suffruticosa and Daphne odora were determined. Phylogenetic trees and pairwise comparisons using complete RNA1- and RNA2-encoded polyproteins showed that the two CNSV isolates are divergent (83.19%-89.42% in polyprotein 1 and 73.61%-85.78% in polyprotein 2). A comparative analysis based on taxonomic criteria for the species demarcation of nepoviruses confirmed that they are not new species but distinct variants. This is the first report of the complete genome sequences of CNSV detected in P. suffruticosa and D. odora, and the first report of CNSV infecting P. suffruticosa.