Use of a three-dimensional microarray system for detection of levofloxacin resistance and the mec A gene in Staphylococcus aureus.
We evaluated a novel three-dimensional microarray (Pam Chip microarray) system to detect the presence of levofloxacin-related resistance mutations and the mec A gene. The results were compared to those obtained for 27 Staphylococcus aureus isolates by conventional DNA sequencing or PCR methods. Hybridization and fluorescence detection were performed using an FD 10 system designed for Pam Chip microarray under conditions optimized for each target/probe on the array. In dilution series analysis using multiplex PCR samples, the sensitivity of the microarray was about 10 times greater than that of conventional PCR methods. A high level of data reproducibility was also confirmed in those analyses. Various point mutations in quinolone resistance-determining regions detected by our system corresponded perfectly to the results obtained by conventional DNA sequencing. The results of the mec A gene detection using our system also corresponded to the PCR method; that is, signal/band was detected in all isolates of methicillin-resistant S. aureus, and no signal/band was detected in any isolate of methicillin-susceptible S. aureus. In conclusion, our novel three-dimensional microarray system provided rapid, specific, easy, and reproducible results for the simultaneous detection of levofloxacin resistance and the mec A gene in S. aureus.
Nagaoka
Tomonori
T
Department of Laboratory Medicine, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Hamamatsu 431-3192, Japan.
Horii
Toshinobu
T
Satoh
Takatomo
T
Ito
Tomoko
T
Monji
Akio
A
Takeshita
Akihiro
A
Maekawa
Masato
M
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics*; D024881:Drug Resistance, Bacterial; D006801:Humans; D064704:Levofloxacin; D015242:Ofloxacin / Q000494:pharmacology*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D046915:Penicillin-Binding Proteins; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D013211:Staphylococcus aureus / Q000145:classification / Q000187:drug effects* / Q000235:genetics
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D046915:Penicillin-Binding Proteins; C487789:mecA protein, Staphylococcus aureus; D064704:Levofloxacin; D015242:Ofloxacin