title: Development of an endpoint genotyping assay to detect the Mycoplasma pneumoniae 23S rRNA gene and distinguish the existence of macrolide resistance-associated mutations at position 2063.
abstract: The prevalence of macrolide-resistant Mycoplasma pneumoniae harboring a mutation in the 23S rRNA gene is increasing, and rapid detection assays are needed for clinical management. We developed an endpoint genotyping assay to detect the M. pneumoniae 23S rRNA gene and determine the existence of macrolide resistance-associated mutations at position 2063 (A2063G, A2063T and A2063C mutations). This A2063B genotyping assay detected more than 50 copies/reaction of the M. pneumoniae gene in every nucleotide mutation at position 2063. Of 42 clinical specimens, 3 were positive without mutation, 6 were positive with the A2063G mutation, and 33 were negative. The results were confirmed using nested PCR with the sequencing of the M. pneumoniae 23S rRNA gene, and a high sensitivity (90%), specificity (100%), and coincidence ratio (kappa coefficient=0.93) were obtained. Therefore, the A2063B genotyping assay is useful for the rapid discrimination of macrolide resistance mutations at position 2063.
authors: Department of Microbiology, Yamagata Prefectural Institute of Public Health, Yamagata 990-0031, Japan; Department of Infectious Diseases, Yamagata University Faculty of Medicine, Yamagata 990-9585, Japan. Electronic address: suzukiyu1@pref.yamagata.jp.
authors: Yu
authors: Y
authors: Suzuki
authors: Department of Microbiology, Yamagata Prefectural Institute of Public Health, Yamagata 990-0031, Japan.
authors: Junji
authors: J
authors: Seto
authors: Department of Infectious Diseases, Yamagata University Faculty of Medicine, Yamagata 990-9585, Japan.
authors: Yoshitaka
authors: Y
authors: Shimotai
authors: Department of Microbiology, Yamagata Prefectural Institute of Public Health, Yamagata 990-0031, Japan.
authors: Tatsuya
authors: T
authors: Ikeda
authors: Yamagata Prefectural Nairiku Meat Inspection Center, Yamagata 990-0892, Japan.
authors: Kazue
authors: K
authors: Yahagi
authors: Department of Microbiology, Yamagata Prefectural Institute of Public Health, Yamagata 990-0031, Japan.
authors: Katsumi
authors: K
authors: Mizuta
authors: Department of Infectious Diseases, Yamagata University Faculty of Medicine, Yamagata 990-9585, Japan.
authors: Yoko
authors: Y
authors: Matsuzaki
authors: Department of Infectious Diseases, Yamagata University Faculty of Medicine, Yamagata 990-9585, Japan.
authors: Seiji
authors: S
authors: Hongo
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D023381:Endpoint Determination / Q000379:methods*; D005838:Genotype; D006801:Humans; D018942:Macrolides / Q000494:pharmacology*; D009154:Mutation; D009177:Mycoplasma pneumoniae / Q000235:genetics* / Q000472:pathogenicity; D011019:Pneumonia, Mycoplasma / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods; D015995:Prevalence; D015347:RNA Probes; D012329:RNA, Bacterial / Q000235:genetics; D012338:RNA, Ribosomal, 23S / Q000235:genetics* / Q000302:isolation & purification*; D012680:Sensitivity and Specificity
chemical_list: D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D018942:Macrolides; D015347:RNA Probes; D012329:RNA, Bacterial; D012338:RNA, Ribosomal, 23S
keywords: Detection; Endpoint genotyping assay; Macrolide resistance; Mycoplasma pneumoniae