title: Development and evaluation of a loop-mediated isothermal amplification assay for rapid and sensitive detection of Vibrio parahaemolyticus.
abstract: Loop-mediated isothermal amplification (LAMP) assays targeting the rpoD and toxR genes were developed to detect Vibrio parahaemolyticus. All 78 tested V. parahaemolyticus strains yielded positive results within 40 min, while negative results were obtained for 69 strains of other organisms even at 60 min. For V. parahaemolyticus ATCC 17802 in pure culture, the detection limits of LAMP assays targeting rpoD and toxR were 3.7 and 450 CFU per test, respectively. Due to the higher sensitivity of rpoD-LAMP, it was further evaluated for the ability to detect V. parahaemolyticus in seafood samples. V. parahaemolyticus populations spiked in short-necked clams were enumerated by the most-probable-number (MPN) method combined with the rpoD-LAMP assay and the MPN method with a culture method using agar medium. The MPN-rpoD-LAMP method had better sensitivity and was more rapid than the conventional method. These results indicate that the MPN-LAMP assay targeting the rpoD gene is a specific, sensitive, and rapid method to enumerate V. parahaemolyticus organisms.
authors: Eiken Chemical Company Ltd., 143 Nogi, Nogi-machi, Shimotsuga-gun, Tochigi 329-0114, Japan. Jiro_Nemoto@eiken.co.jp
authors: Jiro
authors: J
authors: Nemoto
authors: Masanari
authors: M
authors: Ikedo
authors: Tadashi
authors: T
authors: Kojima
authors: Takayoshi
authors: T
authors: Momoda
authors: Hirotaka
authors: H
authors: Konuma
authors: Yukiko
authors: Y
authors: Hara-Kudo
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics*; D015373:Bacterial Typing Techniques; D004268:DNA-Binding Proteins / Q000235:genetics; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D006460:Hemolysin Proteins / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D016133:Polymerase Chain Reaction; D017747:Seafood / Q000382:microbiology*; D012680:Sensitivity and Specificity; D012758:Shellfish / Q000382:microbiology*; D014157:Transcription Factors / Q000235:genetics; D014735:Vibrio Infections / Q000382:microbiology; D014736:Vibrio parahaemolyticus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D006460:Hemolysin Proteins; D014157:Transcription Factors; C066719:toxR protein, bacteria