versión On-line ISSN 1851-7617
Rev. argent. microbiol. v.42 n.2 Ciudad Autónoma de Buenos Aires abr./jun. 2010
Tn7::In2-8 dispersion in multidrug resistant isolates of Acinetobacter baumannii from Chile
1Departamento de Microbiología, Parasitología e Inmunología, Facultad de Medicina, Universidad de Buenos Aires, Buenos Aires, Argentina;
2Facultad de Ciencias Biológicas, Departamento de Microbiología, Laboratorio de Antibióticos, Universidad de Concepción, Concepción, Chile;
3Cátedra de Microbiología, Instituto de Química Biológica, Facultad de Ciencias - Universidad de la República, Montevideo, Uruguay.
*Correspondence. E-mail: firstname.lastname@example.org
Acinetobacter baumannii is considered an important pathogen in our hospital environment having a well-known capacity to acquire different mechanisms of antibiotic resistance. Previous studies in our laboratory had exposed the high dispersion of class 2 integrons in this species. In the present study, we analyzed 7 multiresistant intI2 positive A. baumannii isolates, 6 of which were found to harbour the Tn7::In2-8 element. Our results demonstrate the unusually high distribution of Tn7::In2-8 among different A. baumannii clones from Chile, suggesting a particular behavior of these elements at geographical level.
Key words: Class 2 integron; Acinetobacter baumannii; Antibiotic resistance
Dispersión de Tn7::In2-8 en aislamientos multirresistentes de Acinetobacter baumannii de Chile. Acinetobacter baumannii, patógeno de importancia clínica en el ámbito hospitalario, es reconocido como un microorganismo que posee la capacidad de evolucionar rápidamente hacia la multirresistencia. Estudios previos efectuados en nuestro laboratorio han demostrado la alta dispersión de los integrones de clase 2 en aislamientos de esta especie. En el presente trabajo se analizaron 7 aislamientos de Acinetobacter baumannii multirresistentes portadores de la integrasa de clase 2, 6 de los cuales portaban el inusual arreglo Tn7::In2-8. Nuestros resultados muestran una elevada frecuencia de dispersión del elemento Tn7::In2-8 en diferentes clones circulantes en Chile, lo que sugiere un comportamiento geográfico particular.
Palabras clave: Integrón de clase 2; Acinetobacter baumannii, Resistencia antibiótica
Acinetobacter baumannii (Ab) is considered a serious threat in the hospital environment over the world (2, 5). Recently, it has also emerged as an important pathogen among soldiers returning from the Iraq and Afghanistan war zones (5). Furthermore, some Ab strains have become resistant to almost all currently available antibacterial agents through multifactorial combinatory antimicrobial resistance mechanisms (5). Class 1 and 2 integrons previously found in this species (3, 7) are elements that contain the genetic determinants of the components of a site-specific recombination system which recognizes and captures antimicrobial gene cassettes contributing to the multidrug resistant phenotype (4).
In the GenBank, in contrast with the large amount of cassette rearrangements reported for class 1 integrons, there are only 11 arrays of class 2 integrons described as having a different architecture from the one widespread over the world, embedded in the Tn7 transposon with antimicrobial resistance gene cassettes within the variable region (8). A special characteristic of most type 2 integrase genes described so far is that they are not functional due to the presence of an internal stop codon (7), which most commonly explains the few cassette arrays in this class.
The aim of this study was to characterize 7 intI2 positive. A. baumannii multidrug resistant clinical isolates from 4 distinct regions of Chile, that were randomly chosen from a collection of ninety (n = 90) multiresistant A. baumannii intI2 positive isolates recovered from a total of two hundred and twelve (n = 212) Ab strains from 24 different hospitals (Table 1) during the period 1990-2006. Total DNA was extracted and subjected to PCR amplification with specific primers (7). To detect the inserted gene cassettes, the class 2 integron variable region was amplified, and PCR cartography was also done as previously described (7). Both DNA strands were sequenced, using an ABIPrism 3100 BioAnalyzer equipment. The sequences were analyzed with the Blast V2.0 software (http://www.ncbi.nlm.nih.gov/BLAST/).
Table 1. Description of the clinical isolates used in the study.
Surprisingly, the class 2 integron called Tn7::In2-8 (7), harbouring an unusual array of gene cassettes within the variable region sat2-aadB-catB2(ΔattI2)-dfrA1-sat2-aadA1-orfX-ybfA-ybfB-ybgA was found in 6 out of the 7 isolates. Sequence analysis of the gene cassettes and of unusual cassettes found in the Tn7::In2-8 revealed that all of them are identical to the previously described array (7). The other strain harbored the typically found array in class 2 integrons embedded in the Tn7 transposon. We have previously described the presence of integron Tn7::In2-8 in three out of 106 A. baumannii clinical samples harbouring class 2 integrons (7, 8). The AB1 isolate was recovered in 1994, and isolates AB28 and AB29, in Argentina in the year 2003. Two of the isolates belonged to the same clone, and AB1 belonged to clone IV that was found to have epidemic behaviour in 5 hospitals from Buenos Aires City (1). By repetitive extragenic palindromic DNA sequence-based PCR (REPPCR) and according to the cut-off level previously determined (6), the 7 isolates from this study exhibited 3 distinguishable clones, all different from those detected in Argentina (Table 1). One possibility to explain the high dispersion of class 2 integrons in our A. baumannii isolates, considering these and previous results from our laboratory, is that the different clones of A. baumannii dispersed in the Southern Cone of Latin America are more likely or tend to carry class 2 integrons in their genome (7).
The five transposition genes of the Tn7 transposon were also found in all 7 isolates bearing 100% identity to the transposition genes of accession number X17693. In addition, we determined by PCR and sequence analysis that all Tn7::In2-8 were inserted in the specific attTn7Ab site in the chromosome of the A. baumannii isolates (3), and that all type 2 integrase genes harboured the internal stop codon.
Our results agree with the low amount of cassette arrays in class 2 integrons found over the world. In addition, the putative mechanisms involved in the architecture of gene cassettes in In2-8, homologous recombination and/or illegitimate integrase intermolecular recombination, suggest that recombination mediated by integron integrases is not only playing an important role for array evolution in class 2 integrons. This study also evidences an unusual high distribution of the Tn7::In2-8 element among different A. baumannii clones in 3 different cities of Chile (Valparaíso, Santiago de Chile and Concepción), when compared to the low distribution of class 2 integrons in A. baumannii over the world (8). We have also previously reported the presence of this element in 3 out of 106 intI2 positive isolates of A. baumannii from Argentina (7, 8). This result could correspond to the regional dispersion of Tn7::In2-8 among A. baumannii isolates from Chile and Argentina.
Acknowledgements: M.S.R and D.C. are members of the Carrera del Investigador Científico, CONICET-Argentina. This study was supported by grants M017 from UBACyT and BID/OC 0690 from ANPCyT to D.C., Buenos Aires, Argentina.
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