DNA Fingerprinting of Mycobacterium Tuberculosis: Evaluation of an IS6110-Targeting Fluorescent Amplified Fragment Length Polymorphism DNA Fingerprinting Alternative to IS6110 RFLP

Jason Evans, HPA

Authors: Jason T. Evans, Nicola Thorne, E. Grace Smith, Peter M. Hawkey, Saheer Gharbia, Cath Arnold.

Affiliations: Regional Centre for Mycobacteriology, Health Protection Agency, West Midlands Public Health Laboratory, Birmingham Heartlands Hospital, Birmingham, UK. Applied and Functional Genomics, Centre for Infections, Health Protection Agency, London.

Objectives: A rapid, simple, highly-discriminatory DNA fingerprinting method that produces data that can be easily analysed is the ultimate goal for control of tuberculosis. The current `gold standard', IS6110 RFLP, is far from meeting all these requirements.

Methods: A panel of 78 blinded isolates were selected for analysis by IS6110 RFLP and FAFLP. A novel method called Fluorescent Amplified Fragment Length Polymorphism (FAFLP) that uses capillary electrophoresis to analyse the same genetic element as RFLP was compared with IS6110 RFLP. IS6110 RFLP was carried out using a standardised method.

Results: IS6110 RFLP defined the 78 isolates into nine clusters, with 63 clustered and 15 unique isolates. FAFLP fragment profiles for five repeated digestions of strain H37Rv was 100% reproducible. FAFLP allocated 97% of RFLP-clustered isolates to the same eight clusters as RFLP.

Conclusion: The discriminatory power exhibited by FALP in this initial collection of isolates indicates that it would be a rapid and specific screening alternative to IS6110 RFLP. FAFLP will enable rapid screening of strains for epidemiological investigations, which will direct the prevention of transmission of tuberculosis.