title: Discrimination of Mycobacterium abscessus subsp. massiliense from Mycobacterium abscessus subsp. abscessus in clinical isolates by multiplex PCR.
abstract: The rapidly growing mycobacterium M. abscessus sensu lato is the causative agent of emerging pulmonary and skin diseases and of infections following cosmetic surgery and postsurgical procedures. M. abscessus sensu lato can be divided into at least three subspecies: M. abscessus subsp. abscessus, M. abscessus subsp. massiliense, and M. abscessus subsp. bolletii. Clinical isolates of rapidly growing mycobacteria were previously identified as M. abscessus by DNA-DNA hybridization. More than 30% of these 117 clinical isolates were differentiated as M. abscessus subsp. massiliense using combinations of multilocus genotyping analyses. A much more cost-effective technique to distinguish M. abscessus subsp. massiliense from M. abscessus subsp. abscessus, a multiplex PCR assay, was developed using the whole-genome sequence of M. abscessus subsp. massiliense JCM15300 as a reference. Several primer sets were designed for single PCR to discriminate between the strains based on amplicons of different sizes. Two of these single-PCR target sites were chosen for development of the multiplex PCR assay. Multiplex PCR was successful in distinguishing clinical isolates of M. abscessus subsp. massiliense from samples previously identified as M. abscessus. This approach, which spans whole-genome sequencing and clinical diagnosis, will facilitate the acquisition of more-precise information about bacterial genomes, aid in the choice of more relevant therapies, and promote the advancement of novel discrimination and differential diagnostic assays.
authors.affiliation: Leprosy Research Center, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Kazue
authors.initials: K
authors.lastname: Nakanaga
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Sekizuka
authors.forename: Hanako
authors.initials: H
authors.lastname: Fukano
authors.forename: Yumi
authors.initials: Y
authors.lastname: Sakakibara
authors.forename: Fumihiko
authors.initials: F
authors.lastname: Takeuchi
authors.forename: Shinpei
authors.initials: S
authors.lastname: Wada
authors.forename: Norihisa
authors.initials: N
authors.lastname: Ishii
authors.forename: Masahiko
authors.initials: M
authors.lastname: Makino
authors.forename: Makoto
authors.initials: M
authors.lastname: Kuroda
authors.forename: Yoshihiko
authors.initials: Y
authors.lastname: Hoshino
mesh_terms: D001431:Bacteriological Techniques / Q000379:methods*; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D016680:Genome, Bacterial; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D008969:Molecular Sequence Data; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D009161:Mycobacterium / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D009165:Mycobacterium Infections, Nontuberculous / Q000382:microbiology*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012141:Respiratory Tract Infections / Q000382:microbiology; D017422:Sequence Analysis, DNA; D017192:Skin Diseases, Bacterial / Q000382:microbiology
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S