title: Development of loop-mediated isothermal amplification assays for genotyping of Type III Secretion System in Pseudomonas aeruginosa.
abstract: Pseudomonas aeruginosa is a well-known environmental bacterium capable of causing a variety of life-threatening human infections, with a Type III Secretion System (T3SS) as the most significant virulence determinant. P. aeruginosa strains exhibit unique T3SS virulence genotypes defined by the presence of either exoS or exoU. In this study, loop-mediated isothermal amplification (LAMP) assays for rapid detection of exoS and exoU in P. aeruginosa have been developed and evaluated. Set of four primers were designed for LAMP-based amplification of exoS and exoU respectively. The LAMP reactions were performed at 63°C for 40 min, with detection limits of 100 fg purified DNA. In 107 river water isolates, exoS and exoU were detected in 10 (9%) and 89 (83%) isolates, respectively, and in 38 soil isolates, they were detected in 7 (18%) and 31 (82%) cases respectively. In conclusion, the LAMP assays are rapid, simple and cost-effective tools for detection of the exoU- and exoS-types of P. aeruginosa strains.
  
  
  CONCLUSIONS
  This method can be used for the rapid, sensitive and low-cost detection of genes (exoS and exoU) encoding proteins that are part of Type III Secretion System of Pseudomonas aeruginosa. It can serve as an efficient method in outbreak situations or in routine surveillance studies to judge virulence potential and to investigate pathogenesis of P. aeruginosa.
authors.affiliation: College of Food Science, Southwest University, Chongqing, China.
authors.forename: H
authors.initials: H
authors.lastname: Shi
authors.affiliation: College of Food Science, Southwest University, Chongqing, China.
authors.forename: Z
authors.initials: Z
authors.lastname: Chen
authors.affiliation: College of Food Science, Southwest University, Chongqing, China.
authors.forename: J
authors.initials: J
authors.lastname: Kan
mesh_terms: D036002:ADP Ribose Transferases / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001427:Bacterial Toxins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D005838:Genotype; D060005:Genotyping Techniques; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D011552:Pseudomonas Infections / Q000382:microbiology; D011550:Pseudomonas aeruginosa / Q000235:genetics* / Q000302:isolation & purification / Q000472:pathogenicity; D000069296:Type III Secretion Systems / Q000235:genetics*; D037521:Virulence Factors / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D017931:DNA Primers; D000069296:Type III Secretion Systems; D037521:Virulence Factors; C111749:pseudomonas exoprotein A protein, Pseudomonas aeruginosa; D036002:ADP Ribose Transferases; C444820:exoenzyme S
keywords: Pseudomonas aeruginosa; environment; exoS; exoU; loop-mediated isothermal amplification