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Antimicrobial Agents and Chemotherapy, August 2001, p. 2287-2298, Vol. 45, No. 8
0066-4804/01/$04.00+0   DOI: 10.1128/AAC.45.8.2287-2298.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Novel Class A beta -Lactamase Sed-1 from Citrobacter sedlakii: Genetic Diversity of beta -Lactamases within the Citrobacter Genus

Stephanie Petrella,1 Dominique Clermont,2 Isabelle Casin,3 Vincent Jarlier,1 and Wladimir Sougakoff1,*

Laboratoire de Recherche Moléculaire sur les Antibiotiques, Faculté de Médecine Pitié-Salpêtrière, Université Pierre et Marie Curie,1 Collection de l'Institut Pasteur, Institut Pasteur,2 and Service de Microbiologie, Hôpital Saint-Louis, Université Paris 7,3 Paris, France

Received 20 November 2000/Returned for modification 15 March 2001/Accepted 26 May 2001

Citrobacter sedlakii 2596, a clinical strain resistant to aminopenicillins, carboxypenicillins, and early cephalosporins such as cephalothin, but remaining susceptible to acylureidopenicillins, carbapenems, and later cephalosporins such as cefotaxime, was isolated from the bile of a patient treated with beta -lactam and quinolone antibiotics. The isolate produced an inducible class A beta -lactamase of pI 8.6, named Sed-1, which was purified. Characterized by a molecular mass of 30 kDa, Sed-1 preferentially hydrolyzed benzylpenicillin, cephalothin, and cloxacillin. The corresponding gene, blaSed-1, was cloned and sequenced. Its deduced amino acid sequence shared more than 60% identity with the chromosome-encoded beta -lactamases from Citrobacter koseri (formerly C. diversus) (84%), Klebsiella oxytoca (74%), Serratia fonticola (67%), and Proteus vulgaris (63%) and 71% identity with the plasmid-mediated enzyme MEN-1. A gene coding for a LysR transcriptional regulator was found upstream from blaSed-1. This regulator, named SedR, displayed 90% identity with the AmpR sequence of the chromosomal beta -lactamase from C. koseri and 63 and 50% identity with the AmpR sequences of P. vulgaris and Enterobacter cloacae, respectively. By using DNA-DNA hybridization, a blaSed-1-like gene was identified in two reference strains, C. sedlakii (CIP-105037) and Citrobacter rodentium (CIP-104675), but not in the 18 strains of C. koseri studied. Two DNA fragments were amplified and sequenced from the reference strains of C. sedlakii CIP-105037 and C. rodentium CIP-104675 using two primers specific for blaSed-1. They shared 98 and 80% identity with blaSed-1, respectively, confirming the diversity of the chromosomally encoded class A beta -lactamases found in Citrobacter.

* Corresponding author. Mailing address: L.R.M.A., Laboratoire de Recherche Moléculaire sur les Antibiotiques, Faculté de Médecine Pitié-Salpêtrière, 91 boulevard de l'hôpital, F-75634 Paris Cedex 13, France. Phone: 33 (1) 40 77 97 46. Fax: 33 (1) 45 82 75 77. E-mail: sougakof{at}lmcp.jussieu.fr

Antimicrobial Agents and Chemotherapy, August 2001, p. 2287-2298, Vol. 45, No. 8
0066-4804/01/$04.00+0   DOI: 10.1128/AAC.45.8.2287-2298.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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