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Journal of Clinical Microbiology, November 2005, p. 5622-5627, Vol. 43, No. 11
0095-1137/05/$08.00+0     doi:10.1128/JCM.43.11.5622-5627.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Fast Ligation-Mediated PCR, a Fast and Reliable Method for IS6110-Based Typing of Mycobacterium tuberculosis Complex

Florian Reisig,^1, Kristin Kremer,² Beate Amthor,¹ Dick van Soolingen,² and Walter H. Haas^1,3*

Children's Hospital, University of Heidelberg, Heidelberg, Germany,¹ National Institute of Public Health and the Environment (RIVM), Bilthoven, The Netherlands,² Department for Infectious Disease Epidemiology, Robert Koch Institute, Berlin, Germany³

Received 6 April 2005/ Returned for modification 9 June 2005/ Accepted 9 August 2005

IS6110 restriction fragment length polymorphism (RFLP) analysis is the most widely applied method for strain differentiation of Mycobacterium tuberculosis complex. We have previously described mixed-linker PCR, an IS6110-based PCR method that favorably compared with other typing methods for M. tuberculosis complex according to reproducibility and ability to differentiate between strains. Here we report the further development of this method, called fast ligation-mediated PCR (FLiP), which allows analysis of strains within one working day and starting from less than 1 ng of mycobacterial DNA or a crude cell lysate. Blinded analysis of a standard set of 131 M. tuberculosis complex and nontuberculous isolates showed the ability to differentiate 81 types among 90 M. tuberculosis complex isolates with 84 different IS6110 RFLP fingerprint patterns and detected 97% of the 31 duplicate samples. We suggest that FLiP can serve to rapidly detect chains of transmission prior to starting high-throughput analysis or standard IS6110 RFLP. It may as well serve as a secondary typing technique for other, non-IS6110-based methods.

Present address: Abteilung für Anästhesie und Intensivmedizin, BG Unfallklinik, Murnau, Germany.

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