Asclepias yellow vein virus: a proposed new potyvirus in milkweed in the USA

In February 2018, foliar chlorosis and vein yellowing symptoms (Fig. 1) were observed in a nursery Bloomington, Minnesota, USA Asclepias curassavica plants grown from stem cuttings originating Houston, Texas. Filamentous virus-like particles 12 nm diameter length ranging approximately 750–800 by transmission electron microscopy (TEM) negatively stained partially purified leaf tissue extracts (Ahlawat et al., 1996) diseased 2), but not asymptomatic seed. No other extracts. To determine the identity of filamentous virus associated with A. curassavica, nucleic acid was isolated phenol-chloroform extraction virions extract. Based on approximate size detected TEM, group-specific primers designed for detection Carlavirus Potyvirus species used to whether viruses either group present (Chen 2002). A single PCR product 1.6 kb using potyvirus primers, no obtained carlavirus primers. The products cloned pGEM®-T Easy Vector System (Promega, USA) five clones sequenced Sanger sequencing both directions. BLAST search showed highest 89% at nucleotide level sequence deposited as (GenBank Accession No. MH898481) syriaca USA. next most closely related Shallot yellow stripe (DQ925458) Clover isolate P180 (D89544), 71% identity. partial GenBank (MH809364). According demarcation criteria potyviruses, optimal threshold coat protein is 76–77% level. these results, we propose that described this study are isolates new within genus. Beyond sequence, further information about has been published. name (AsYVV) proposed describe virus. role AsYVV disease Asclepias, ten swamp milkweed (A. incarnata) seed determined be virus-free TEM reported above, then mechanically inoculated crude extract infected tissue. Leaf similar those occurring original plant growth starting two months post-inoculation 3). Confirmation presence symptomatic after inoculation done RT-PCR. visualisation revealed flexuous particles. RT-PCR confirmation Illustra Ready-To-Go Beads (GE Healthcare Life Sciences, USA). primer pair sequence; F2 (5'-ACTAGGTCAACTCAAGATCAAGC-3') R2 (5'-TTCTCTAGCTCAATAGAATAGAATACC-3'). temperature cycling programme consisted reverse-transcription reaction 42°C 45 min, enzyme inactivation 95°C 5 min followed 30 cycles 94°C s, annealing 57°C s 72°C final extension time min. Products expected (867 bp) plants. amplicons before. Ten directions resulting sequences shared high (98-99%) initial (MH898481). experiments confirmed mechanical AsYVV. This first report infecting incarnata. previously syriaca. Visualisation performed Phillips CM12 Transmission Electron Microscope assistance Gail Celio University Minnesota - Imaging Centers.