title: An assessment of a multiplex PCR assay for differentiating clinically important mycobacteria based on pncA gene variation.
abstract: The pncA genes in mycobacteria are responsible for the production of pyrazinamidase (PZase). In Mycobacterium tuberculosis, PZase hydrolyses pyrazinamide (PZA) to pyrazonic acid, a compound that possesses bactericidal activity against tubercle bacilli. Nucleotide sequences of pncA genes found within mycobacteria where aligned in an effort to ascertain the significance of any variability in sequence. Three sets of primers (one degenerate and five consensus sequences) were designed and employed in a multiplex PCR assay to amplify the pncA region in seven clinically common mycobacteria. The banding patterns generated from each species in conjunction with PZase activity tests demonstrated that the mycobacterial species examined could be clearly identified and differentiated from one another. Although not yet tested with clinical isolates, the combination of these two assays has provided a promising discriminatory tool for the identification of commonly encountered clinical mycobacteria species.
authors.affiliation: Department of Infectious Diseases and Microbiology, Faculty of Medicine, Imperial College, Du Cane Road, London, W12 0NN, UK. m.bamaga@ic.ac.uk
authors.forename: Mohammad S
authors.initials: MS
authors.lastname: Bamaga
authors.forename: David J
authors.initials: DJ
authors.lastname: Wright
authors.forename: Hongyi
authors.initials: H
authors.lastname: Zhang
mesh_terms: D000581:Amidohydrolases / Q000235:genetics*; D015373:Bacterial Typing Techniques / Q000379:methods*; D001483:Base Sequence; D017931:DNA Primers; D004587:Electrophoresis, Agar Gel; D014644:Genetic Variation; D009161:Mycobacterium / Q000235:genetics* / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D016415:Sequence Alignment
chemical_list: D017931:DNA Primers; D000581:Amidohydrolases; C022490:pyrazinamide deamidase