title: Identification and Differentiation of Monilinia Species Causing Brown Rot of Pome and Stone Fruit using High-Resolution Melting (HRM) Analysis.
abstract: Brown rot is a devastating disease of stone fruit caused by Monilinia spp. Among these species, Monilinia fructicola is a quarantine pathogen in Europe but has recently been detected in several European countries. Identification of brown rot agents relies on morphological differences or use of molecular methods requiring fungal isolation. The current study was initiated to develop and validate a high-resolution melting (HRM) method for the identification of the Monilinia spp. and for the detection of M. fructicola among other brown rot pathogens. Based on the sequence of the cytb intron from M. laxa, M. fructicola, M. fructigena, M. mumecola, M. linhartiana, and M. yunnanensis isolates originating from several countries, a pair of universal primers for species identification and a pair of primers specific to M. fructicola were designed. The specificity of the primers was verified to ensure against cross-reaction with other fungal species. The melting curve analysis using the universal primers generated six different HRM curve profiles, each one specific for each species. Τhe HRM analysis primers specific to M. fructicola amplified a 120-bp region with a distinct melt profile corresponding to the presence of M. fructicola, regardless of the presence of other species. HRM analysis can be a useful tool for rapid identification and differentiation of the six Monilinia spp. using a single primer pair. This novel assay has the potential for simultaneous identification and differentiation of the closely related Monilinia spp. as well as for the differentiation of M. fructicola from other common pathogens or saprophytes that may occur on the diseased fruit.
authors.affiliation: First and third authors: Plant Pathology Laboratory, Faculty of Agriculture, Forestry and National Resources, Thessaloniki, Aristotle University of Thessaloniki, Greece; second author: Institute of Applied Biotechnology, CERTH, Thermi, Thessaloniki, Greece.
authors.forename: Antonios
authors.initials: A
authors.lastname: Papavasileiou
authors.affiliation: First and third authors: Plant Pathology Laboratory, Faculty of Agriculture, Forestry and National Resources, Thessaloniki, Aristotle University of Thessaloniki, Greece; second author: Institute of Applied Biotechnology, CERTH, Thermi, Thessaloniki, Greece.
authors.forename: Panagiotis B
authors.initials: PB
authors.lastname: Madesis
authors.affiliation: First and third authors: Plant Pathology Laboratory, Faculty of Agriculture, Forestry and National Resources, Thessaloniki, Aristotle University of Thessaloniki, Greece; second author: Institute of Applied Biotechnology, CERTH, Thermi, Thessaloniki, Greece.
authors.forename: George S
authors.initials: GS
authors.lastname: Karaoglanidis
mesh_terms: D001203:Ascomycota / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D002681:China; D017931:DNA Primers / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics / Q000302:isolation & purification; D005638:Fruit / Q000382:microbiology*; D006115:Greece; D010935:Plant Diseases / Q000382:microbiology*; D016133:Polymerase Chain Reaction; D000068239:Prunus persica / Q000382:microbiology*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013030:Spain; D044366:Transition Temperature; D014481:United States
chemical_list: D017931:DNA Primers; D004271:DNA, Fungal
keywords: peach