A Mixed Infection of Helenium Virus S With Two Distinct Isolates of Butterbur Mosaic Virus, One of Which Has a Major Deletion in an Essential Gene

John Hammond, Michael Reinsel, Samuel Grinstead, Ben Lockhart, Ramon Jordan, Dimitre Mollov

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Multiple carlaviruses infect various ornamental plants, often having limited host ranges and causing minor symptoms, yet often reducing yield or quality. In this study we have identified a mixed infection of butterbur mosaic virus (ButMV) and helenium virus S (HelVS) from a plant of veronica (Veronica sp.) showing foliar mosaic and distortion. Carlavirus-like particles were observed by transmission electron microscopy (TEM), and RNA from partially purified virions was amplified by random RT-PCR, yielding clones of 439–1,385 bp. Two partially overlapping clones including coat protein (CP) sequence, and two of four partial replicase clones, were closely related to ButMV-J (AB517596), previously reported only from butterbur (Petasites japonicus) in Japan. Two other partial replicase clones showed lower identity to multiple carlaviruses. Generic primers which amplify the 3′-terminal region of multiple carlaviruses yielded clones of three distinct sequences: (1) with 98% nt identity to HelVS; (2) ButMV-A, showing 82% nt identity to ButMV-J; and (3) ButMV-B, with 78% nt identity to each of ButMV-J and ButMV-A. Further amplification of upstream fragments revealed that ButMV-B had an internal deletion in TGB1, confirmed using isolate-specific primers. Near-complete genomes of both ButMV-A and ButMV-B were obtained from next-generation sequencing (NGS), confirming the deletion within ButMV-B, which is presumably maintained through complementation by ButMV-A. HelVS was previously reported only from Helenium hybrids and Impatiens holstii. A near-complete HelVS genome was obtained for the first time by NGS from the same sample. Additional Veronica hybrids infected with HelVS were identified by TEM and RT-PCR, including cv. ‘Sunny Border Blue’ which was also subjected to NGS. This resulted in assembly of an 8,615 nt near-complete HelVS genome, with high identity to that from the mixed infection. The predicted CP sequence has 96% amino acid (aa) identity to HelVS from helenium (Q00556). Other ORFs show a maximum of 54% (TGB3) to 68% (NABP) aa identity to the equivalent ORFs of other carlaviruses. These results demonstrate for the first time maintenance by complementation of a carlavirus isolate with a major deletion in an essential gene, and confirm that HelVS is a distinct species in the genus Carlavirus.

Original languageEnglish (US)
Article number612936
JournalFrontiers in Microbiology
StatePublished - Dec 21 2020

Bibliographical note

Funding Information:
This work was supported by internal USDA-ARS funding under projects 8020-22000-042-00D and 8020-22000-302-00D.

Publisher Copyright:
© Copyright © 2020 Hammond, Reinsel, Grinstead, Lockhart, Jordan and Mollov.


  • carlavirus
  • complementation
  • defective isolate
  • next-generation sequencing
  • veronica


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