title: DNA sequencing by Microseq kit targeting 16S rRNA gene for species level identification of mycobacteria.
abstract: OBJECTIVE
  Identification of mycobacteria to the species level is of therapeutic significance. Conventional methods are laborious and time consuming so we did 16S rRNA sequencing using a commercial MicroSeq sequencing kit, which includes DNA sequencing with software package for identification and phylogenetic analysis of clinical mycobacterial isolates.
  
  
  METHODS
  A total of 47 mycobacteria were tested by both conventional and genotypic method using commercially available MicroSeq 500 amplification kit assay. The identification was determined by comparing the 500 bp amplified product of 16S rDNA sequence to the MicroSeq database.
  
  
  RESULTS
  The phenotypic identification was concordant with genotypic identification in 33 (70.2%) isolates of 14 Mycobacterium tuberculosis, 11 M. fortuitum, 7 M. abscessus and 1 M. duvalii. For the discrepant isolates, identification was possible only by DNA sequencing in 14 (29.7%) isolates. The 14 discrepant isolates were 5 M. farcinogenes, 3 M. genavense, 2 M. species. nov and 1 each of M. fortuitum, M. immuogenum, M. simiae and M. wolinskyi. Of these, five were uncommon species that were difficult to identify by phenotypic method.
  
  
  CONCLUSIONS
  The MicroSeq DNA sequencing is an excellent tool for species identification of mycobacteria, which reduces the turn around time, makes repeat analysis easy as compared to phenotypic identification specially for mycobacterial isolates with ambiguous biochemical profiles.
authors.affiliation: L & T Microbiology Research Centre, Vision Research Foundation, Sankara Nethralaya, Chennai, India. drklt@snmail.org
authors.forename: K Lily
authors.initials: KL
authors.lastname: Therese
authors.forename: John
authors.initials: J
authors.lastname: Bartell
authors.forename: P
authors.initials: P
authors.lastname: Deepa
authors.forename: S
authors.initials: S
authors.lastname: Mangaiyarkarasi
authors.forename: Diedra
authors.initials: D
authors.lastname: Ward
authors.forename: Joseph
authors.initials: J
authors.lastname: Dajcs
authors.forename: H N
authors.initials: HN
authors.lastname: Madhavan
authors.forename: David
authors.initials: D
authors.lastname: Stroman
mesh_terms: D017931:DNA Primers / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D005838:Genotype; D009159:Mycobacteriaceae / Q000145:classification / Q000166:cytology / Q000235:genetics*; D010641:Phenotype; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods; D013045:Species Specificity
chemical_list: D017931:DNA Primers; D021903:DNA, Ribosomal Spacer; D012336:RNA, Ribosomal, 16S