Development of nevel genotyping method of Pseudomonas aeruginosa and clinical application

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We developed a novel digital genotyping method of Pseudomonas aeruginosa (P. aeruginosa PCR-based ORF typing method; POT typing method). According to the distribution of the ORF (Open reading frame) in the clinical isolates, 10 ORF originated from genomic islet and 5 ORF originated from genomic island to increase the discriminatory power were selected. Metallo-beta-lactamase (MBL) (IMP or VIM group) gene and P. aeruginosa marker gene were added for the typing, finally the primers were adapted for the 10 and 9-plex PCR. The PCR results were scored with a 1 or 0 according to the presence or absence of the amplicon of the selected genes in order of primer number, and the resulting binary code was transformed into decimal numbers, separating into two parts with 10 (genomic islet, POT1) and 9 (genomic island and MBL, POT2) figures. POT1 was shown to be related with Multilocus Sequence Typing (MLST) number, and the discriminatory power was equivalent to PFGE.