title: Development and use of a multiplex polymerase chain reaction assay based on Apx toxin genes for genotyping of Actinobacillus pleuropneumoniae isolates.
abstract: Actinobacillus pleuropneumoniae (A. pleuropneumoniae) is the etiological agent of a porcine pleuropneumonia that threatens the global swine industry. The major pathogenic toxins of A. pleuropneumoniae include ApxI, ApxII, ApxIII, and ApxIV, which are serotype or serovar specific. Several techniques have been developed for the identification and typing of A. pleuropneumoniae. Serological assays are used to identify and serotype A. pleuropneumoniae, but factors such as cross-reactivity limit their specificity. Labor, time, and the requirement for specific antibodies are also drawbacks of these assays. Multistep polymerase chain reaction (PCR) techniques based on apx genes have been reported for the identification and typing of A. pleuropneumoniae. This study developed multiplex PCR for the identification and genotyping of A. pleuropneumoniae based on apx genes. This multiplex PCR technique was successful in differentiating 11 of 15 reference serotypes. Five different primer sets were used to amplify the 4 apx genes from each serotype in a single-step reaction. The multiplex PCR reported in this study was further used in genotyping 51 field isolates of A. pleuropneumoniae from different regions of Korea. The concomitant amplification of all 4 apx genes makes multiplex PCR more specific and convenient for the diagnosis and genotyping of A. pleuropneumoniae.
authors.affiliation: Department of Infectious Diseases, College of Veterinary Medicine, School of Agricultural Biotechnology, Seoul National University, Seoul 151-742, Korea.
authors.forename: Nabin
authors.initials: N
authors.lastname: Rayamajhi
authors.forename: Sung Jae
authors.initials: SJ
authors.lastname: Shin
authors.forename: Sang Gyun
authors.initials: SG
authors.lastname: Kang
authors.forename: Deog Yong
authors.initials: DY
authors.lastname: Lee
authors.forename: Jeong Min
authors.initials: JM
authors.lastname: Ahn
authors.forename: Han Sang
authors.initials: HS
authors.lastname: Yoo
mesh_terms: D000189:Actinobacillus Infections / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary; D016977:Actinobacillus pleuropneumoniae / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D001427:Bacterial Toxins / Q000235:genetics*; D005838:Genotype; D006460:Hemolysin Proteins; D016133:Polymerase Chain Reaction / Q000379:methods*; D012703:Serotyping / Q000662:veterinary; D013552:Swine; D013553:Swine Diseases / Q000175:diagnosis / Q000382:microbiology
chemical_list: C085361:ApxII toxin, bacteria; C079721:ApxIII toxin protein, Actinobacillus; C122177:ApxIVA protein, Actinobacillus pleuropneumoniae; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D006460:Hemolysin Proteins; C087744:ApxI toxin, Bacteria