title: Direct detection of isoniazid-resistant Mycobacterium tuberculosis in respiratory specimens by multiplex allele-specific polymerase chain reaction.
abstract: This study evaluated the feasibility of using 2 multiplex allele-specific polymerase chain reaction (MAS-PCR) assays targeting 2 mutations (codon 315 of the katG gene and the 15th nucleotide preceding the mabA-inhA operon) to directly detect isoniazid (INH)-resistant Mycobacterium tuberculosis in cultured isolates and respiratory specimens. A total of 203 M. tuberculosis isolates and 487 respiratory specimens were investigated. The MAS-PCR assays successfully amplified all M. tuberculosis isolates and acid-fast bacilli smear-positive specimens while only 49.2% of the smear-negative specimens exhibited positive MAS-PCR results. The MAS-PCR assays identified 83.4% and 79.2% of the resistant strains in the culture isolates and respiratory specimens, respectively. All the inferred genotypes were in complete accordance with subsequent DNA sequence analyses. This study suggested the application of our improved MAS-PCR protocols to provide the rapid identification of INH-resistant M. tuberculosis directly in respiratory specimens. The technical simplicity, short turnaround time, and low cost of this molecular strategy should facilitate routine diagnostic services in developing areas with a high prevalence of drug-resistant tuberculosis.
authors.affiliation: Department of Microbiology, Queen Mary Hospital, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong Special Administrative Region, China.
authors.forename: Gilman Kit Hang
authors.initials: GK
authors.lastname: Siu
authors.forename: Yuk Ho
authors.initials: YH
authors.lastname: Tam
authors.forename: Pak Leung
authors.initials: PL
authors.lastname: Ho
authors.forename: Ann Siew Gek
authors.initials: AS
authors.lastname: Lee
authors.forename: Tak Lun
authors.initials: TL
authors.lastname: Que
authors.forename: Cindy Wing Sze
authors.initials: CW
authors.lastname: Tse
authors.forename: Kam Tong
authors.initials: KT
authors.lastname: Yip
authors.forename: Jason Tsz Hin
authors.initials: JT
authors.lastname: Lam
authors.forename: Vincent Chi Chung
authors.initials: VC
authors.lastname: Cheng
authors.forename: Kwok Yung
authors.initials: KY
authors.lastname: Yuen
authors.forename: Wing Cheong
authors.initials: WC
authors.lastname: Yam
mesh_terms: D000483:Alleles; D019943:Amino Acid Substitution / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D002374:Catalase / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial; D006801:Humans; D007538:Isoniazid / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests / Q000379:methods; D020125:Mutation, Missense; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics* / Q000302:isolation & purification; D010088:Oxidoreductases / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D013183:Sputum / Q000382:microbiology*; D013997:Time Factors; D014376:Tuberculosis / Q000382:microbiology*
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D010088:Oxidoreductases; D002374:Catalase; C480353:katG protein, Mycobacterium tuberculosis; C085257:InhA protein, Mycobacterium; D007538:Isoniazid