Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/16/2009
Publication Date: 7/10/2009
Publication URL: http://hdl.handle.net/10113/45119
Citation: Vaira, A.M., Hansen, M.A., Murphy, C., Reinsel, M.D., and Hammond, J. 2009. First report of Freesia sneak virus in Freesia sp. in Virginia, USA. Plant Disease. 93(9):965. Available http://dx.doi.org/10.1094/PDIS-93-9-0965B. Interpretive Summary: Previously un-reported virus-like diseases are periodically observed in ornamental crops. The causal agents of these disease must be identified in order to allow detection and identification of the source of the disease, and if possible, the elimination of the virus from planting stocks. Leaf necrosis disease symptoms were observed in a commercial freesia planting in Virginia in 2008, and studies were carried out to identify the causal agent. Electron microscopy, and molecular detection methods utilizing the polymerase chain reaction (PCR) were used to identify the presence of an ophiovirus in most of the affected plants. Cloning and sequencing of the viral coat protein gene allowed identification of the ophiovirus as Freesia sneak virus, previously reported only from Europe and South Africa. Electron microscopy also revealed the presence of flexuous particles typical of potyviruses; PCR assays using primers specific for potyviruses yielded an amplification product of the expected size from only 2 of 15 plants tested. Sequence analysis identified the potyvirus as Freesia mosaic virus. This was the first time that Freesia sneak virus has been detected in freesias in the USA; identification of the virus, and availability of a method for its detection will allow growers the option of screening planting stocks for presence of the disease agent prior to planting.
Technical Abstract: In spring 2008 freesias (cvs. ‘Honeymoon’ and ‘Santana’) with striking virus-like symptoms were received by the Virginia Tech Plant Disease Clinic from a cut flower nursery in Gloucester, VA, and forwarded to the USDA-ARS Floral and Nursery Plants Research Unit in Beltsville, MD, for analysis. About 25% of the plants had coalescing interveinal chlorotic, whitish necrotic, or dark brown to purple necrotic spots on leaves (leaf necrosis syndrome). Symptomatic plants were scattered within the planting. Fifteen symptomatic plants collected between March and May 2008 were analyzed for ophiovirus infection by RT-PCR using ophiovirus-specific degenerate primers. The diagnostic 136 bp ophiovirus product from the RdRp gene was amplified from 14 out of 15 freesia plants tested. A partially purified preparation was analyzed by transmission electron microscopy; potyvirus- as well as ophiovirus-like particles were detected. RT-PCR was also performed on the same samples using potyvirus-specific degenerate primers D335 and U335; the diagnostic 335 bp fragment was amplified from 2 out of 15 plants; the sequence from these plants was identified as Freesia mosaic virus (FreMV). The ophiovirus coat protein (CP) gene was amplified by RT-PCR from an extract of two symptomatic freesia plants using primers designed from the sequence of an Italian Freesia sneak virus (FreSV) isolate (GenBank accession DQ885455) from freesia. The c.1.3 kb expected amplified fragment was cloned using the TOPO vector system (Invitrogen) and two clones were sequenced on both strands. The deduced amino acid sequences showed 99% identity with the Italian FreSV CP sequence, confirming the presence of FreSV in the symptomatic freesia plants. This is the first report of FreSV in both Virginia and the United States. Soil-borne Freesia leaf necrosis disease has been reported in Europe since the 1970’s; several viral causal agents have been hypothesized but recent findings correlate best with the ophiovirus. A varicosavirus (Freesia leaf necrosis virus) and unknown agents may also be involved as Koch’s postulates for the necrotic syndrome have not been fulfilled for any agent. In Virginia the presence of FreSV, but not FreMV, was strongly correlated with the leaf necrosis syndrome. FreSV, likely soil-borne through Olpidium brassicae, may pose a new soil-borne threat for bulbous ornamentals, as it has been recently detected also in Lachenalia spp. (Hyacinthaceae) from South Africa. Although specific testing for O. brassicae was not performed, the disease may potentially persist in soil for years in O. brassicae resting spores and development of symptoms may be affected by environmental conditions.