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Antimicrobial Agents and Chemotherapy, October 2005, p. 4418-4420, Vol. 49, No. 10
0066-4804/05/$08.00+0     doi:10.1128/AAC.49.10.4418-4420.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

LETTER TO THE EDITOR

Class 2 Integron with a Novel Cassette Array in a Burkholderia cenocepacia Isolate


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LETTER
 
Burkholderia cepacia complex (BCC) organisms are gram-negative opportunistic emerging pathogens associated with a poor prognosis for patients with cystic fibrosis (CF) (10). Carbapenems and ceftazidime are administered to patients suffering from BCC infections, and trimethoprim-sulfamethoxazole has historically been the drug of choice (6). Recently, the acquisition of resistance determinants to sulfamethoxazole located in the 3' conserved region of class 1 integrons in BCC isolates from CF patients has been described (2).

The goal of this study was to determine the presence of class 1, 2, and 3 integrons in a Burkholderia cenocepacia strain (BC1) isolated from the sputum of a 14-year-old CF patient in a surgery and transplant center in Buenos Aires, Argentina.

The phenotypic analysis was performed using biochemical reactions (4) and led us to identify the isolate as BCC. The genotypic identification by the recA PCR-restriction fragment length polymorphism method (9) resulted in a IIIB recA lineage, corresponding to Burkholderia cenocepacia. A search for the presence of the cable pilin subunit gene (cblA) and the B. cepacia epidemic strain marker, by PCR with specific primers (9), yielded negative results. The BC1 strain was susceptible to trimethoprim-sulfamethoxazole (MIC = 1 µg/ml), meropenem (MIC = 0.012 µg/ml), ceftazidime (MIC = 0.19 µg/ml), and minocycline (MIC = 1 µg/ml) according to CLSI (formerly NCCLS) (1, 7).

By analysis of the class 1, 2, and 3 integrons present in the BC1 strain by PCR with specific primers (Table 1), we found only a class 2 integron (8). In order to identify the inserted gene cassettes, PCR cartography was performed with primers described in Table 1. Two PCR products, obtained with the 125'CS and satR primers and the Inti2R and satR primers (Fig. 1) were sequenced on both strands using an ABI 373 sequencer and analyzed using the Genetics Computer Group (GCG) software (Wisconsin Package, version 10.3). Only a sat2 gene cassette was found in the variable region, and it is noteworthy that this novel rearrangement did not have the orfX cassette always present in Tn7-like transposons (Fig. 1). Also, the analysis of the sequence led us to determine that the internal stop codon in the class 2 integrase gene was present as it has been previously described (3). Therefore, this novel rearrangement could likely have arisen from the action of a class 1 integrase from another element in trans (3). We also detected the presence of tnsE and tnsD genes located in the 3' conserved sequence region and involved in Tn7 transposition (11) (Table 1 and Fig. 1). We have named it Tn7::In2-1 (accession number DQ082896) in order to identify novel rearrangements of class 2 integrons in a simple manner.


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  		TABLE 1. Primers used in this study



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  		FIG. 1. Variable region of the class 2 integrons of the Tn7 family described in the literature, Tn7 (accession number NC_002525), Tn1825 (X56815), Tn4132 (Z50804), Tn7::IS1-ereA (AY183453), a recently published class 2 integron with accession number AB161461, and Tn7::In2-1 from this study. The thin vertical open bar represents the attI2 site, the ovals represent the attC sites of the gene cassettes, and dashed lines mean that the sequence is not available. All intI2 genes sequenced reveal an internal stop codon (3).

Our results provide evidence that class 2 integrons located in the Tn7 family of transposons are able to be incorporated in the genome of BCC, as described for class 1 integrons recently (2).


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ACKNOWLEDGMENTS
 
This study was supported by a grant from UBACYT MO17, Buenos Aires, Argentina, to D.C.


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REFERENCES
 
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  1. Clinical and Laboratory Standards Institute. 2005. Performance standards for antimicrobial susceptibility testing, 14th informational supplement. CLSI document M100-S15. Clinical and Laboratory Standards Institute, Wayne, Pa.
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  2. Crowley, D., M. Daly, B. Lucey, P. Shine, J. J. Collins, B. Cryan, J. E. Moore, P. Murphy, G. Buckley, and S. Flanning. 2002. Molecular epidemiology of cystic fibrosis-linked Burkholderia cepacia complex isolates from three national referral centres in Ireland. J. Appl. Microbiol. 92:992-1004.[CrossRef][Medline]
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  3. Hansson, K., L. Sundstrom, A. Pelletier, and P. Roy. 2002. IntI2 integron integrase in Tn7. J. Bacteriol. 184:1712-1721.[Abstract/Free Full Text]
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  6. Lewin, C., C. Doherty, and J. Govan. 1993. In vitro activities of meropenem, PD 127391, PD 131628, ceftazidime, chloramphenicol, co-trimoxazole, and ciprofloxacin against Pseudomonas cepacia. Antimicrob. Agents Chemother. 37:123-125.[Abstract/Free Full Text]
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  8. Orman, B. E., S. A. Piñeiro, S. Arduino, M. Galas, R. Melano, M. I. Caffer, D. O. Sordelli, and D. Centrón. 2002. Evolution of multiresistance in nontyphoid Salmonella serovars from 1984 to 1998 in Argentina. Antimicrob. Agents Chemother. 46:3963-3970.[Abstract/Free Full Text]
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  9. Petrucca, A., P. Cipriani, P. Valenti, D. Santapaola, C. Cimmino, G. L. Scoarughi, I. Santino, S. Stefani, R. Sessa, and M. Nicoletti. 2003. Molecular characterization of Burkholderia cepacia isolates from cystic fibrosis (CF) patients in an Italian CF center. Res. Microbiol. 154:491-498.[Medline]
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  10. Urban, T. A., A. Griffith, A. M. Torok, M. E. Smolkin, J. L. Burns, and J. B. Golberg. 2004. Contribution of Burkholderia cenocepacia flagella to infectivity and inflammation. Infect. Immun. 72:5126-5134.[Abstract/Free Full Text]
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  11. Waddell, C. S., and N. L. Craig. 1988. Tn7 transposition: two transposition pathways directed by five Tn7-encoded genes. Genes Dev. 2:137-149.[Abstract/Free Full Text]
María Soledad Ramírez
Liliana Jordá Vargas
Departamento de Microbiología, Parasitología e Inmunología
Facultad de Medicina
Universidad de Buenos Aires
Buenos Aires, Argentina,1

Viviana Cagnoni
Marta Tokumoto
Laboratorio de Microbiología
ICYCC
Fundación Favaloro
Buenos Aires, Argentina,2

Daniela Centrón*
Departamento de Microbiología, Parasitología e Inmunología
Facultad de Medicina
Universidad de Buenos Aires
Paraguay 2155, P-12, Capital Federal, Argentina,3

* Phone: 54 11 5950-9500, ext. 2171, Fax: 54 11 4964 2554, E-mail: dcentron{at}fmed.uba.ar

Antimicrobial Agents and Chemotherapy, October 2005, p. 4418-4420, Vol. 49, No. 10
0066-4804/05/$08.00+0     doi:10.1128/AAC.49.10.4418-4420.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

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  • Parks, A. R., Peters, J. E. (2007). Transposon Tn7 Is Widespread in Diverse Bacteria and Forms Genomic Islands. J. Bacteriol. 189: 2170-2173 [Abstract] [Full Text]  
  • Barlow, R. S., Gobius, K. S. (2006). Diverse class 2 integrons in bacteria from beef cattle sources. J Antimicrob Chemother 58: 1133-1138 [Abstract] [Full Text]  

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