Molecular typing of Legionella pneumophila serogroup 1 by pulsed-field gelelectrophoresis with SfiI and comparison of this method with restriction fragment-length polymorphism analysis

A panel of 131 well-characterised Legionella pneumophila serogroup 1 strain s, previously used to evaluate a restriction fragment-length polymorphisms (RFLP) typing scheme, was examined by pulsed-held gel electrophoresis (PFGE ) with the restriction endonuclease SfiI, The data obtained show that PFGE with SfiI is highly discriminatory method yielding an index of discriminati on (IOD) of 0.992 and 0.975, with 100% and 90% similarity thresholds respec tively, compared with an IOD of 0.909 for the RFLP typing method. Reproduci bility of PFGE profiles within gels was excellent and it was possible to co mpare the profiles visually. However, the reproducibility of the technique between gels was poor and visual comparison of the patterns was extremely d ifficult. Computer-aided analysis assisted the assessment of inter-gel repr oducibility, Of 11 duplicates examined only four pairs showed 100% similari ty, although 9 of 11 showed greater than or equal to 90% similarity. In an attempt to determine if the PFGE banding patterns were sufficiently unambig uous to allow the method to be used as a definitive typing method, 20 coded strains were examined. At a 90% similarity level, 16 of these were placed in the correct PFGE type and four were not allocated to a type. Partial dig estion of DNA by SfiI was noted despite careful control of DNA and enzyme c oncentrations, suggesting that an alternative enzyme might give more reprod ucible results.