title: Assessment of fluorescent amplified fragment length polymorphism analysis for epidemiological genotyping of Legionella pneumophila serogroup 1.
abstract: This study assessed the reproducibility and epidemiological concordance of double-enzyme fluorescent amplified fragment length polymorphism (fAFLP) analysis for genotyping of Legionella pneumophila serogroup (sg) 1. fAFLP fragment analysis was performed on three different sequencing platforms (one gel- and two capillary-based) in different laboratories with a well-characterised set of 50 strains of L. pneumophila sg 1. fAFLP data were analysed with the Pearson correlation similarity coefficient, using a range of parameters, and dendrogram outputs were converted to arbitrary types after selection of a specified percentage similarity threshold. The results obtained were compared with those obtained by the standard non-fluorescent AFLP method and were found to be broadly concordant. Using optimised settings for each fAFLP method to analyse the panel of 50 strains, epidemiological concordance (E) and reproducibility (R) values of 1.00 were obtained, and the number of types ranged from nine to 15, compared with E=1.00 and R=1.00, with 16 types, for the non-fluorescent AFLP protocol. The study demonstrated the potential of fAFLP for typing strains of L. pneumophila sg 1 on all three platforms; however, inter-platform comparison of fAFLP data was not achieved. fAFLP analysis may have a role in the fingerprinting of multiple isolates during Legionella outbreak investigations, but further work is required before type designations and identification libraries can be developed.
authors.affiliation: Health Protection Agency, Respiratory & Systemic Infection Laboratory, Centre for Infections, London, UK. Norman.Fry@HPA.org.uk
authors.forename: N K
authors.initials: NK
authors.lastname: Fry
authors.forename: B
authors.initials: B
authors.lastname: Afshar
authors.forename: P
authors.initials: P
authors.lastname: Visca
authors.forename: D
authors.initials: D
authors.lastname: Jonas
authors.forename: J
authors.initials: J
authors.lastname: Duncan
authors.forename: E
authors.initials: E
authors.lastname: Nebuloso
authors.forename: A
authors.initials: A
authors.lastname: Underwood
authors.forename: T G
authors.initials: TG
authors.lastname: Harrison
mesh_terms: D016172:DNA Fingerprinting / Q000379:methods*; D004269:DNA, Bacterial / Q000235:genetics; D005060:Europe / Q000453:epidemiology; D005453:Fluorescence; D005838:Genotype; D006801:Humans; D016952:Legionella pneumophila / Q000235:genetics; D007877:Legionnaires' Disease / Q000453:epidemiology*; D017720:Molecular Epidemiology / Q000379:methods*; D015203:Reproducibility of Results
chemical_list: D004269:DNA, Bacterial