Mixedlinker polymerase chain reaction: a new method for rapid fingerprinting of isolates of the Mycobacterium tuberculosis complex

Rapid recognition of multidrug-resistant strains of Mycobacterium tuberculosis is a desirable goal for treatment of patients and protection of health care workers. DNA fingerprints produced with the insertion sequence IS6110 generate restriction fragment length polymorphism (RFLP) patterns that reliably identify M. tuberculosis complex strains. This report describes a rapid technique for RFLP typing using the polymerase chain reaction. The method uses one primer specific for IS6110 and a second primer complementary to a linker ligated to the restricted genomic DNA. In one strand the linker contains uracil in place of thymidine, and specific amplification is obtained by elimination of this strand with uracil N-glycosylase. Mixed-linker fingerprinting clearly differentiated multidrug-resistant isolates from 12 outbreaks and unambiguously assigned them to 26 RFLP groups. Tuberculosis has made a dramatic comeback in industri-alized countries, in part because of the AIDS pandemic (17). In addition, the treatment of tuberculosis has been compli-cated by the appearance of drug-resistant strains spreading readily in the hospital setting (29). Since 1990 an increasing number of nosocomial outbreaks, especially among AID