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Antimicrobial Agents and Chemotherapy, April 2001, p. 1022-1029, Vol. 45, No. 4
0066-4804/01/$04.00+0   DOI: 10.1128/AAC.45.4.1022-1029.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Rapid Screening Technique for Class 1 Integrons in Enterobacteriaceae and Nonfermenting Gram-Negative Bacteria and Its Use in Molecular Epidemiology

Alison J. Maguire,dagger Derek F. J. Brown, James J. Gray, and Ulrich Desselberger*

Clinical Microbiology & Public Health Laboratory, Addenbrooke's Hospital, Cambridge, United Kingdom.

Received 2 May 2000/Returned for modification 23 August 2000/Accepted 19 December 2000

A screening technique for integrons in members of the family Enterobacteriaceae and nonfermenting gram-negative bacteria by real-time PCR is reported. A total of 226 isolates of gram-negative bacteria obtained from a variety of clinical specimens were screened for class 1 integrons by real-time PCR performed on a LightCycler instrument. This technique used a primer pair specific for a 300-bp conserved region at the 5' ends of class 1 integrons. The screening assay was evaluated by comparison with results obtained by the conventional, thermal-block PCR (long PCR) by using established conditions and primers for the detection of class 1 integrons, and the real-time PCR technique was thus shown to be both sensitive and specific. DNA from 50 of 226 (22%) isolates screened was identified as containing an integron by the screening PCR, and sequence data were obtained across the integron for 34 of 50 (68%) of these isolates. In an attempt to study the molecular epidemiology of antimicrobial resistance genes carried within integrons, a comparison of the types of gene cassettes carried by isolates from different patients was made. Adenyltransferase genes conferring resistance to streptomycin and spectinomycin were the predominant gene cassettes amplified in the study. Resistance to trimethoprim was also frequently found to be encoded within integrons. Furthermore, multiple bacterial isolates obtained from one patient over a 5-month period were all shown to carry an integron containing the same single adenyltransferase gene cassette, suggesting that these elements were relatively stable in this case.

* Corresponding author. Mailing address: Clinical Microbiology & Public Health Laboratory, Box 236, Addenbrooke's Hospital, Hills Road, Cambridge CB2 2QW, United Kingdom. Phone: 44 1223 216816. Fax: 44 1223 242775. E-mail: ulrich.desselberger{at}msexc.addenbrookes.anglox.nhs.uk.

dagger Present address: Department of Medicine, University of Cambridge, Addenbrooke's Hospital, Cambridge, United Kingdom.

Antimicrobial Agents and Chemotherapy, April 2001, p. 1022-1029, Vol. 45, No. 4
0066-4804/01/$04.00+0   DOI: 10.1128/AAC.45.4.1022-1029.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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