First Report of Rose Rosette Virus Associated with Rose Rosette Disease Affecting Roses in California
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Rose rosette disease (RRD) affecting rose (Rosa spp.) was first described in the western United States and Canada in the early 1940s (Conners 1941) and is now widely distributed in the eastern United States. Typical RRD symptoms include excessive lateral shoot growth and thorniness, witches broom, leaf proliferation and malformation, mosaic, red pigmentation, and eventual plant decline and death. RRD is caused by the negative-sense RNA virus, rose rosette virus (RRV), which is vectored by the eriophyid mite (Phyllocoptes fructiphilus Keifer) (Di Bello et al. 2015; Laney et al. 2011). California is one of the main producers of roses in the United States, with a wholesale value of US$17.6 million in 2016 (NASS 2016). In August 2017, symptoms suggestive of RRD were observed in a commercial nursery in Wasco, CA. Leaf tissues were collected from a total of 415 plants, representing many rose cultivars, throughout the entire nursey. Two plants (cvs. Veterans Honor and Brilliant Pink Iceberg) showed RRD symptoms, whereas the rest of the samples were asymptomatic. Total RNA was extracted from leaf petioles using the MagMA-96 viral RNA isolation kit (Thermo Fisher Scientific, Waltham, MA) and tested for RRV using a modified quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay developed by Dobhal et al. (2016). Only the two symptomatic plants were positive for RRV. Plants were also microscopically examined, and the P. fructiphilus was observed on several plants throughout the nursery. In June 2018, three garden roses (unknown cultivars) planted in two neighboring homes in Bakersfield, CA, approximately 50 km from the 2017 finds, were identified with RRD symptoms. Leaves from all three plants tested positive for RRV by RT-qPCR and were also infested with P. fructiphilus. To confirm these results, total nucleic acid was extracted from stored (80C) leaves from each of the five symptomatic plants and used for cDNA library construction after a ribosomal RNA depletion step (TruSeq Stranded Total RNA with RiboZero Plant kit, Illumina, San Diego, CA). High-throughput sequencing (HTS) was performed on the Illumina NextSeq 500 platform, generating between 15 million and 41 million 75-nucleotide (nt) single-end raw data reads per sample. Analysis of the HTS reads identified nearly complete sequences (>97%) of the seven RRV genome segments from all five samples. The RRV genomes from the two nursery plants sampled in 2017 shared 100% nt identity to one another. The RRV genomes from the three homeowner plants sampled in 2018 shared 100% nt identity to one another but not to the nursery samples (nursery versus homeowner showed 98% nt identity). Sequences from the two locations shared 98% nt identity to the NCBI reference RRV RNA3 (NC_015300) and RNA7 (NC_034981) and 99% nt identity to the NCBI reference RRV RNA1-2 (NC_015298-99) and RNA4-6 (NC_015301, NC_034979, and NC_034980). These results suggest that the samples from these two locations are likely two separate introductions of RRV into California. Two genomic sequences, one from each location, were submitted to GenBank under the accession numbers MH581213 to MH581226. To our knowledge, this is the first report of RRV associated with RRD in California. Further studies will be necessary to assess the relevance and potential threat of this virus to the garden rose nursery industry in California.
