Comment[ArrayExpressAccession] E-GEOD-43629 MAGE-TAB Version 1.1 Public Release Date 2013-07-17 Investigation Title New rapid scheme for distinguishing subspecies of the Mycobacterium abscessus group and identification of Mycobacterium massiliense with inducible clarithromycin resistance Comment[Submitted Name] New rapid scheme for distinguishing subspecies of the Mycobacterium abscessus group and identification of Mycobacterium massiliense with inducible clarithromycin resistance Experiment Description Mycobacterium abscessus [M. abscessus (sensu lato) or M. abscessus group] comprises three closely related taxa with taxonomic status under revision: M. abscessus sensu stricto, M. bolletii and M. massiliense. We describe here a simple, robust and cost effective PCR-based method for distinguishing among M. abscessus, M. massiliense and bolletii. Based on the M. abscessus ATCC 19977T genome, discriminatory regions were identified between M. abscessus and M. massiliense from array-based comparative genomic hybridization. A typing scheme using PCR primers designed for four of these locations was applied to 46 well-characterized clinical isolates comprising 29 M. abscessus, 15 M. massiliense and 2 M. bolletii previously identified by multi-target sequencing. Interestingly, 2 isolates unequivocally identified as M. massiliense were shown to have a full length erm(41) instead of the expected gene deletion and showed inducible clarithromycin resistance after 14 days. We propose using this PCR-based typing scheme combined with erm(41) PCR for a straightforward identification of M. abscessus, M. massiliense and M. bolletii and assessment of inducible clarithromycin resistance. This method can be easily implemented into a routine workflow providing subspecies level identification within 24 hours of isolation of M. abscessus group. Two-color CGH with 4 independent Mycobacterium clinical isolates and the M massiliense type strain (CCUG 48898) labeled with Cy3 were cohybridized with the M abscessus type strain (ATCC 19977) labeled with Cy5 on a tiling array designed against the M abscessus type strain Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Myers Shallom Gardina Myers Sebastian Conville Tettelin Olivier Uzel Sampaio Holland Zelazny Person First Name Timothy Shamira Paul Timothy Yinong Patricia Hervé Kenneth Gulbu Elizabeth Steven Adrian Person Mid Initials G J J G N P M M Person Email niaid-mrf-geo@nih.gov Person Affiliation National Institute of Allergy and Infectious Diseases Person Address Research Technologies Branch, National Institute of Allergy and Infectious Diseases, 50 South Drive, Room 5509, Bethesda, MD, USA Person Roles submitter Protocol Name P-GSE43629-1 P-GSE43629-3 P-GSE43629-4 P-GSE43629-2 P-GSE43629-5 Protocol Description Signal distributions were median normalized between arrays, then the ratios (Cy3/Cy5) from the duplicate tiling probes were averaged ID_REF = VALUE = normalized log2 ratios (Cy3/Cy5) [Cy5 used for common reference sample] Direct labeling using Agilent Genomic DNA Enzymatic Labeling Kit Agilent Oligo aCGH Hybridization Kit Lysozyme, CTAB/NaCL, SDS/proteinase K/ chloroform isoamyl alcohol/isopropanol/70% ethanol wash/dry pellet/dissolve in TE Agilent microarray scanner at 3 micron resolution using XDR of 100% and 10% PMT. Agilent Feature Extraction software version 10.7.3.1, protocol CGH_107_Sep09 Protocol Type normalization data transformation protocol labelling protocol hybridization protocol nucleic acid extraction protocol array scanning protocol Experimental Factor Name CLATHROMYCIN RESISTANCE SOURCE ISOLATION DATE ORGANISM Experimental Factor Type clathromycin resistance source isolation date organism Comment[SecondaryAccession] GSE43629 Comment[GEOReleaseDate] 2013-07-17 Comment[ArrayExpressSubmissionDate] 2013-01-18 Comment[GEOLastUpdateDate] 2013-07-20 Comment[AEExperimentType] comparative genomic hybridization by array SDRF File E-GEOD-43629.sdrf.txt