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Journal of Clinical Microbiology, November 1998, p. 3133-3137, Vol. 36, No. 11
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Fluorescent Amplified-Fragment Length Polymorphism Analysis of an Outbreak of Group A Streptococcal Invasive Disease

Meeta Desai,1 Asha Tanna,2 Robert Wall,3 Androulla Efstratiou,2 Robert George,2 and John Stanley1,*

Molecular Biology Unit1 and Respiratory and Systemic Infection Laboratory,2 Central Public Health Laboratory, London NW9 5HT, and Department of Microbiology, Northwick Park Hospital, Harrow, Middlesex,3 United Kingdom

Received 13 May 1998/Returned for modification 9 July 1998/Accepted 2 August 1998

Fluorescent amplified-fragment length polymorphism (FAFLP) analysis was carried out for an outbreak of group A streptococcal (GAS) invasive disease. Streptococcal genomic DNAs were digested with endonucleases EcoRI and MseI, site-specific adaptors were ligated, and PCR amplification was carried out with an EcoRI adaptor-specific primer labelled with fluorescent dye. Amplified fragments of up to 600 bp in size were separated on a polyacrylamide sequencing gel which contained internal size markers in each lane. These data were automatically scanned and analyzed, fragments were precisely sized (±1 bp), and electropherograms were generated for each genome with GeneScan 2.1 software. All isolates were compared in this way. Among 27 GAS isolates examined, we found 18 FAFLP profiles, compared with 12 macrorestriction profiles by pulsed-field gel electrophoresis. FAFLP readily distinguished genotypes for two clones of GAS serotype M77 which were responsible for outbreaks of invasive disease in a care-of-the-elderly system. It provided an automated analysis of the whole genome of bacterial isolates. It was reproducible, more discriminatory, and capable of higher throughput than other molecular typing methods. Given agreed conditions, FAFLP would be reproducible between laboratories for rapid characterization of outbreak strains.

* Corresponding author. Mailing address: Molecular Biology Unit, Central Public Health Laboratory, 61 Colindale Ave., London NW9 5HT, United Kingdom. Phone: 0181 200 4400, ext. 3071. Fax: 0181 200 1569. E-mail: mdesai{at}hgmp.mrc.ac.uk.

Journal of Clinical Microbiology, November 1998, p. 3133-3137, Vol. 36, No. 11
0095-1137/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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