title: Rapid and sensitive detection of Little cherry virus 2 using isothermal reverse transcription-recombinase polymerase amplification.
abstract: Little cherry virus 2 (LChV2) (genus Ampelovirus) is the primary causal agent of little cherry disease (LCD) in sweet cherry (Prunus avium) in North America and other parts of the world. This mealybug-transmitted virus does not induce significant foliar symptoms in most sweet cherry cultivars, but does cause virus-infected trees to yield unevenly ripened small fruits with poor flavor. Most fruits from infected trees are unmarketable. In the present study, an isothermal reverse transcription-recombinase polymerase amplification (RT-RPA) technique was developed using LChV2 coat protein specific primers and probe. Detection of terminally labeled amplicons was achieved with a high affinity lateral flow strip. The RT-RPA is confirmed to be simple, fast, and specific. In comparison, although it retains the sensitivity of RT-PCR, it is a more cost-effective procedure. RT-RPA will be a very useful tool for detecting LChV2 from crude extracts in any growth stage of sweet cherry from field samples.
authors.affiliation: Department of Plant Pathology, Washington State University, IAREC, 24106 North Bunn Road, Prosser, WA 99360, USA.
authors.forename: Tefera A
authors.initials: TA
authors.lastname: Mekuria
authors.affiliation: Agdia, Inc., 30380 County Road 6, Elkhart, IN 46514, USA.
authors.forename: Shulu
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Department of Plant Pathology, Washington State University, IAREC, 24106 North Bunn Road, Prosser, WA 99360, USA. Electronic address: keastwell@wsu.edu.
authors.forename: Kenneth C
authors.initials: KC
authors.lastname: Eastwell
mesh_terms: D001483:Base Sequence; D029381:Closteroviridae / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers / Q000235:genetics; D010935:Plant Diseases / Q000821:virology*; D000068220:Prunus avium / Q000821:virology*; D012367:RNA, Viral / Q000235:genetics; D045522:Recombinases; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013997:Time Factors
chemical_list: D017931:DNA Primers; D012367:RNA, Viral; D045522:Recombinases
keywords: Isothermal detection; Little cherry disease; Little cherry virus 2; Prunus avium; Recombinase polymerase amplification; Sweet cherry