Smilax auriculata: A new host for Orchid fleck virus identified in Florida, USA

Authors: DEY, KISHORE KUMAR - Florida Department Of Agriculture And Consumer Services; VILEZ-CLIMENT, MARIA - Florida Department Of Agriculture And Consumer Services; Padmanabhan, Chellappan; NUNZIATA, SCHYLER - US Department Of Agriculture (USDA); Rivera, Yazmin; MCVAY, JOHN - Florida Department Of Agriculture And Consumer Services; Roy, Avijit

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/12/2021
Publication Date: 6/29/2022
Citation: Dey, K., Vilez-Climent, M., Padmanabhan, C., Nunziata, S., Rivera, Y., Mcvay, J., Roy, A. 2022. Smilax auriculata: A new host for Orchid fleck virus identified in Florida, USA. Plant Disease. https://doi.org/10.1094/PDIS-09-21-2085-PDN.
DOI: https://doi.org/10.1094/PDIS-09-21-2085-PDN

Interpretive Summary: Citrus leprosis (CiL) was first reported from Florida in 1860s but disappeared in mid-1960s. The outbreak in Florida was attributed to infection by an ancestor of Orchid fleck virus (OFV). At present there are four known strains of OFV exist in nature and all of them are either present in Mexico, Colombia, South Africa or in Hawaii and cause CiL. The OFV is transmitted by the flat mite; Brevipalpus californicus. In June 2020, two orchid strains of OFV were identified in three ornamentals, namely Liriope, Aspidistra, and Ophiopogon. Further finding of Smilax as a new host for OFV in Florida increases the vulnerability of other economically important plant species. There is high chance of re-emergence of CiL in Florida as susceptible host, virulent pathogen, appropriate environment, and mite vector are present. Strict preventive measures need to be taken to avoid the re-emergence and establishment of CiL disease in United States.

Technical Abstract: Smilax auriculata is commonly known as Dune/Ear-leaf Greenbrier, belongs to the family Smilacaceae, order Liliales. In June 2020, Orchid fleck virus (OFV) was detected in the Liriope spp. in Florida. In October, S. auriculata showing yellowing and mottling symptoms was tested using OFV specific conventional RT-PCR assay and amplified partial N and L genes. Amplicons sequence shared 99% nucleotide identity with OFV infecting orchids and citrus. The OFV subgroup I (OFV-Orc1) and subgroup II (OFV-Orc2) specific primers were utilized to confirm the presence of OFV type strains infecting Smilax. Sanger sequencing of subgroup I specific amplicons (MZ645934) shared 99% nucleotide identity with OFV-Orc1 (LC222629) whereas subgroup II specific amplicon sequence (MZ645930) shared 98-99 % nucleotide identity with OFV-Orc2 (AB244417). To reconfirm the presence of OFV in Smilax and identify the strain, symptomatic tissue and positive RNA control was sent to USDA-APHIS-PPQ-Beltsville laboratory. Total RNA was extracted from both the samples and tested utilizing optimized conventional RT-PCR protocols with additional sets of OFV generic primers. RT-PCR assays amplified 626 and 831 bp amplicons of P and L gene, respectively, in the symptomatic Smilax samples. The L gene sequence shared 99% identity with both OFV-Orc strains whereas P gene sequence shared 98% nt identity with OFV-Orc2 infecting orchids (AB244417) and citrus (MW021482). A TruSeq Stranded Total RNA Library Prep kit (Illumina) was used to prepare Ribo-depleted RNA library before pursued high throughput sequencing (HTS) on a NextSeq Illumina platform. The resulting 28.2 million 2x75bp reads were de novo assembled using SPAdes v.3.10.0 (Bankevitch et al. 2012). Assembled reads identified 7 non-overlapping viral contigs. Five RNA1 and two RNA2 contigs covered more than 97% of the OFV genome with depth of coverage 5297.61 and 5186.04, respectively. Contigs of RNA1 and RNA2 shared 98-99% nt identity to OFV-Orc2-RNA1 (AB244417) and OFV-Orc-RNA2 (AB244418 and LC222630). No other pathogen sequences were identified. This is the first-time Smilax has been identified as a natural host of OFV. Both the orchid strains are known to infect citrus and cause leprosis in South Africa and Hawaii. Citrus leprosis (CiL) was first reported from Florida in 1860s but disappeared in mid-1960s. The outbreak in Florida was attributed to infection by an ancestor of OFV, citrus leprosis virus-N0. Very recent findings of OFV-Orc in Florida in Liriope, Aspidistra, and Ophiopogon among the Asparagaceae family members and now in the Smilacaceae increases the vulnerability of other economically important plant species. Strict preventive measures need to be taken to avoid the re-emergence of CiL disease in Florida.