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Antimicrobial Agents and Chemotherapy, January 2009, p. 335-336, Vol. 53, No. 1
0066-4804/09/$08.00+0     doi:10.1128/AAC.00584-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

LETTER TO THE EDITOR

High Prevalence of the aac(6')-Ib-cr Gene and Its Dissemination among Enterobacteriaceae Isolates by CTX-M-15 Plasmids in Bulgaria

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Since 1998, three mechanisms of plasmid-mediated quinolone resistance (PMQR) have been reported: Qnr-mediated topoisomerase protection (6), enzymatic modification of ciprofloxacin and norfloxacin by the aminoglycoside acetyltransferase AAC(6')-Ib-cr (10), and active efflux due to QepA (8). PMQR genes confer low-level quinolone resistance and are frequently cotransmitted with extended-spectrum β-lactamase (ESBL) genes (9).

We report the prevalence of aac(6')-Ib-cr and its association with qnr genes in ESBL-producing Enterobacteriaceae isolates in a Bulgarian hospital.

A total of 163 ESBL-producing enterobacteria (4.6% of 3,516 consecutive isolates) were recovered among 10 species at increasing overall prevalence rates between 2000 and 2005 (Table 1). These increases reflected the increasing rates of ESBL production in Escherichia coli (from 1.2% in 2000 to 10.0% in 2005), whereas similar rates of 7% in Klebsiella pneumoniae and the irregular appearance of ESBL production in other species have been observed during the study period. As shown in Table 1, using primers 5'-ACTGAGCATGACCTTGCGATGC-3' and 5'-TTAGGCATCACTGCGTGTTCG-3', aac(6')-Ib was detected in 99 (60.7%) of the ESBL producers distributed among nine species. Of these, 52 (52.5%) were found to carry the cr variant by sequencing, including 2 Citrobacter freundii isolates; one isolate each of Enterobacter aerogenes, Morganella morganii, and K. pneumoniae (all from 2005); and 47 E. coli isolates recovered since 2002, increasing from 0% to 67% during that period. For seven C. freundii isolates, including the two aac(6')-Ib-cr-positive isolates, PCR for qnrA, qnrB, and qnrS (11, 12) yielded amplicons only for qnrB, identified as qnrB10 (GenBank accession number DQ631414), qnrB13 (GenBank accession number EU273755), and qnrB18 (GenBank accession number AM919399) by using sequencing (1). The overall prevalence of qnrB (7/163; 4.3%) was sevenfold lower than that of aac(6')-Ib-cr (52/163, 31.9%), and the coexpression of QnrB and AAC(6')-Ib-cr occurred only in two (1.4% of all) isolates.

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TABLE 1. Distribution of aac(6')-Ib-cr and qnrB genes among 163 ESBL-producing enterobacterial isolates at the National Oncology Center, Sofia, Bulgaria, from 2000 to 2005 and the respective ESBL prevalence rates

The 52 aac(6')-Ib-cr-positive isolates were characterized by antibiotic susceptibility testing and bla content determination as previously described (3, 13). Forty-three isolates were resistant to ciprofloxacin, 41 to gentamicin, and 25 to trimethoprim-sulfamethoxazole. All isolates were resistant to tobramycin and exhibited reduced susceptibility to amikacin, but worrisomely, 39 (75%) of the isolates were classified as amikacin susceptible according to CLSI breakpoints. Fifty isolates had both bla_CTX-M-15 and bla_OXA-1. Of these, 2 isolates also carried bla_SHV-12 and 32 carried bla_TEM-1. The remaining two aac(6')-Ib-cr-positive isolates expressed ESBLs not of the TEM, SHV, CTX-M, VEB, PER, or GES type, associated with the TEM-1 enzyme.

Thirty different XbaI-pulsed-field gel electrophoresis patterns were observed among the 47 E. coli isolates (data not shown). These findings suggest that the high prevalence of aac(6')-Ib-cr was not solely due to the spread of a specific E. coli clone. Transferability of AAC(6')-Ib-cr determinant in broth and on filters was examined using rifampin-resistant recipient E. coli ML4909 (F^– galK2 galT22 hsdR metB1 relA supE44 Rif^r) (4). Transconjugants were selected on bromothymol blue lactose agar containing rifampin (200 µg/ml) and kanamycin (25 µg/ml). Conjugative transfer of aac(6')-Ib-cr was achieved for 42 of the 52 isolates, including one isolate each of K. pneumoniae and M. morganii, two C. freundii isolates, and 38 E. coli isolates. aac(6')-Ib-cr was mostly cotransferred with bla_CTX-M-15 and bla_OXA-1, variably with bla_TEM-1, but not with bla_SHV-12 and qnrB.

This is the first report of qnrB and aac(6')-Ib-cr in clinical Enterobacteriaceae isolates from a Bulgarian hospital. The aac(6')-Ib and its cr variant were highly prevalent in ESBL-producing E. coli. CTX-M-15 plasmid-mediated dissemination of aac(6')-Ib-cr among Enterobacteriaceae isolates was particularly observed, as has been found in other countries (2, 5). In this work, qnrB had a low prevalence and was not cotransferred with the aac(6')-Ib-cr gene. This result supports previous findings suggesting that aac(6')-Ib-cr might already be widespread and substantially more prevalent than qnr genes (7, 10). Most of the isolates carrying the aac(6')-Ib-cr variant were resistant to ciprofloxacin, probably reflecting its ability to promote higher-level quinolone resistance mutations (10).

 
This work was supported by Grant-in-Aid for Scientific Research 17-05764 from the Japan Society for the Promotion of Science.

 
Published ahead of print on 10 November 2008.

Present address: Laboratory for Clinical Microbiology, National Oncology Center, Sofia 1756, Bulgaria.

Top LETTER REFERENCES

 

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						Stefana Sabtcheva* Mitsuo Kaku Department of Infection Control and Laboratory Diagnostics Tohoku University Graduate School of Medicine 1-1 Seiryo-machi Aoba-ku, Sendai 980-8574, Japan Tomoo Saga Yoshikazu Ishii Department of Microbiology and Infectious Diseases Toho University School of Medicine Faculty of Medicine Tokyo, Japan Todor Kantardjiev Department of Microbiology National Center of Infectious and Parasitic Diseases Sofia, Bulgaria
						* Phone: (359) 2 8076293, Fax: (359) 2 8720651, E-mail: stefanasabtcheva{at}gmail.com

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Antimicrobial Agents and Chemotherapy, January 2009, p. 335-336, Vol. 53, No. 1
0066-4804/09/$08.00+0     doi:10.1128/AAC.00584-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

• Strahilevitz, J., Jacoby, G. A., Hooper, D. C., Robicsek, A. (2009). Plasmid-Mediated Quinolone Resistance: a Multifaceted Threat. Clin. Microbiol. Rev. 22: 664-689 [Abstract] [Full Text]  

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