title: Selective detection of viable seed-borne Acidovorax citrulli by real-time PCR with propidium monoazide.
abstract: In recent years, use of the DNA-intercalating dye propidium monoazide (PMA) in real-time PCR has been reported as a novel method to detect viable bacteria in different types of samples, such as food, environmental, and microbiological samples. In this study, viable cells of Acidovorax citrulli, the causal agent of bacterial seedling blight and fruit blotch, were selectively detected and differentiated from dead cells by real-time fluorescent polymerase chain reaction amplification after the bacterial solution was treated with the DNA-binding dye PMA. The primers and TaqMan probe were based on the A. citrulli genome (Aave_1909, Gene ID: 4669443) and were highly specific for A. citrulli. The detection threshold of this assay was 103colony-forming units per mL (CFU/mL) in pure cell suspensions containing viable and dead cells and infected watermelon seeds. Application of this assay enables the selective detection of viable cells of A. citrulli and facilitates monitoring of the pathogen in watermelon and melon seeds.
authors.affiliation: Chinese Academy of Inspection and Quarantine, Beijing 100176, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Tian
authors.affiliation: Shenzhen Entry-Exit Inspection and Quarantine Bureau, Shenzhen 518045, China.|Shenzhen Academy of Inspection and Quarantine, Shenzhen 518010, China.
authors.forename: Jian-Jun
authors.initials: JJ
authors.lastname: Feng
authors.affiliation: Shaanxi University of Technology, Shaanxi 723001, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Hu
authors.affiliation: Chinese Academy of Inspection and Quarantine, Beijing 100176, China.
authors.forename: Wen-Jun
authors.initials: WJ
authors.lastname: Zhao
mesh_terms: D001386:Azides / Q000737:chemistry; D029773:Citrullus / Q000382:microbiology; D042621:Comamonadaceae / Q000235:genetics* / Q000302:isolation & purification; D059022:Food Safety / Q000379:methods*; D011419:Propidium / Q000031:analogs & derivatives / Q000737:chemistry; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012639:Seeds / Q000382:microbiology*
chemical_list: D001386:Azides; C533957:propidium monoazide; D011419:Propidium