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Antimicrobial Agents and Chemotherapy, August 2009, p. 3399-3404, Vol. 53, No. 8
0066-4804/09/$08.00+0 doi:10.1128/AAC.01522-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Multiresistance in Pasteurella multocida Is Mediated by Coexistence of Small Plasmids
Alvaro San Millan,1
Jose Antonio Escudero,1
Belen Gutierrez,1
Laura Hidalgo,1
Nerea Garcia,2
Montserrat Llagostera,3,4
Lucas Dominguez,2 and
Bruno Gonzalez-Zorn1,2*
Departamento de Sanidad Animal, Facultad de Veterinaria, Universidad Complutense de Madrid, Madrid, Spain,1 Centro de Vigilancia Sanitaria Veterinaria (VISAVET), Universidad Complutense de Madrid, Madrid, Spain,2 Departament de Genètica i de Microbiologia, Universitat Autònoma de Barcelona, 08193 Cerdanyola del Vallès (Bellaterra), Barcelona, Spain,3 Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, 08193 Bellaterra, Barcelona, Spain4
Received 14 November 2008/ Returned for modification 1 February 2009/ Accepted 2 June 2009
In most gram-negative bacteria, acquired multiresistance is conferred by large plasmids compiling numerous antimicrobial resistance genes. Here, we show an evolutionary alternative strategy used by Pasteurella multocida to become resistant to multiple clinically relevant antibiotics. Thirteen β-lactam-resistant clinical isolates, concomitantly resistant to tetracyclines and/or streptomycin as well as to sulfonamides, were studied. Pulsed-field gel electrophoresis analysis revealed different profiles among the isolates, showing that clonal dissemination was not the sole event responsible for the spread of multiresistance. Each P. multocida strain carried two or three small plasmids between 4 and 6 kb in size. A direct association between resistance profile and plasmid content was found. Complete nucleotide sequencing of all plasmids revealed seven different replicons, six of them belonging to the ColE1 superfamily. All plasmids carried one, or a maximum of two, antimicrobial resistance determinants. Plasmids pB1000 and pB1002 bore blaROB-1, pB1001 carried tet(B), pB1003 and pB1005 carried sul2 and strA, pB1006 harbored tet(O), and p9956 bore the tet(H) gene. All plasmids except pB1002 and pB1006 were successfully transformed into Escherichia coli. pB1000, also involved in β-lactam resistance in Haemophilus parasuis (A. San Millan et al., Antimicrob. Agents Chemother. 51:2260-2264, 2007), was mobilized in E. coli using the conjugation machinery of an IncP plasmid. Stability experiments proved that pB1000 was stable in P. multocida but highly unstable in E. coli. In conclusion, blaROB-1 is responsible for β-lactam resistance in P. multocida in Spain. Coexistence and the spread of small plasmids are used by P. multocida to become multiresistant.
* Corresponding author. Mailing address: Departamento de Sanidad Animal, Facultad de Veterinaria, Universidad Complutense de Madrid, 28040 Madrid, Spain. Phone: 34 91 3943707. Fax: 34 91 3943908. E-mail: bgzorn{at}vet.ucm.es
Published ahead of print on 15 June 2009.
Antimicrobial Agents and Chemotherapy, August 2009, p. 3399-3404, Vol. 53, No. 8
0066-4804/09/$08.00+0 doi:10.1128/AAC.01522-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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