Variable Regions Of Integrons Research Articles

Detecting Class 1 Integrons and Their Variable Regions in Escherichia coli Whole-Genome Sequences Reported from Andean Community Countries.

Various genetic elements, including integrons, are known to contribute to the development of antimicrobial resistance. Class 1 integrons have been identified in E. coli isolates and are associated with multidrug resistance in countries of the Andean Community. However, detailed information on the gene cassettes located on the variable regions of integrons is lacking. Here, we investigated the presence and diversity of class 1 integrons, using an in silico approach, in 2533 whole-genome sequences obtained from EnteroBase. IntFinder v1.0 revealed that almost one-third of isolates contained these platforms. Integron-bearing isolates were associated with environmental, food, human, and animal origins reported from all countries under scrutiny. Moreover, they were identified in clones known for their pathogenicity or multidrug resistance. Integrons carried cassettes associated with aminoglycoside (aadA), trimethoprim (dfrA), cephalosporin (blaOXA; blaDHA), and fluoroquinolone (aac(6')-Ib-cr; qnrB) resistance. These platforms showed higher diversity and larger numbers than previously reported. Moreover, integrons carrying more than three cassettes in their variable regions were determined. Monitoring the prevalence and diversity of genetic elements is necessary for recognizing emergent patterns of resistance in pathogenic bacteria, especially in countries where various factors are recognized to favor the selection of resistant microorganisms.

Quantification of class 1 integrons and characterization of the associated gene cassettes in the high Arctic – Interplay of humans and glaciers in shaping the aquatic resistome

The Arctic is one of the regions most affected by global climate change, and is subjected to changes linked with a melting cryosphere and increasing anthropopressure. Although antibiotic resistance is a global problem, the diversity and spread of antibiotic-resistant bacteria (ARB), antibiotic resistance genes (ARGs) and integrons in the Arctic are strongly understudied. Therefore, the main aims of this study are the (1) determination of the type and frequency of integron-integrase genes and characterization of incorporated gene cassettes in the genomes of culturable bacteria and (2) quantitative analysis of class 1 integron-integrase gene and human mitochondrial DNA (Hmt-DNA) in the metagenome as markers of anthropogenic impact on the high Arctic environments of the Svalbard Archipelago. Samples of ice, water and sediments were collected in the most populated area of Svalbard, Longyearbyen and its vicinity. Sampling was conducted along an environmental gradient with varying levels of human activity. The environmental gradient started from glaciers, following the proglacial river, the seashore, and the fjord bottom water, including untreated wastewater outflow to the sea. Class 1 integrons were detected in ARB isolated from glacial environments, freshwater and seawater, including wastewater outflow. Moreover, in the variable regions of integrons, genes determining different functions, including antibiotic resistance, virulence and physiological traits were found. These genes play crucial roles in the adaptation of bacteria to cold and dynamic environments. The relative abundance of intI1 genes were reported in metagenomes with different relationships to human activity (ice cores vs wastewater outflow), with the highest mean values observed in the wastewater outflow, and was positively correlated with abundance of the Hmt gene, revealing both natural and human roles in shaping the polar aquatic resistome.

Antimicrobial Resistance and Molecular Characterization of Gene Cassettes from Class 1 Integrons in Escherichia coli Strains.

To investigate the antimicrobial resistance and molecular characterization of gene cassettes from class 1 integrons in Escherichia coli strains isolated from hospitalized patients. Bacterial identification was conducted using the Vitek-2 Compact system, and antimicrobial susceptibility analysis was performed using the Kirby-Bauer method. Class 1 integrons, integrase genes, the variable regions of integrons and promoters from the isolated E. coli were screened by polymerase chain reaction, and subjected to DNA sequencing. In total, 138 E. coli strains were collected from the hospitalized patients, most from urine specimens (41.30%, 57/138). Antimicrobial resistance to ampicillin (89.86%) was most prevalent, with 79.99% of strains being multidrug-resistant (MDR). The class 1 integron integrase intI1 gene was detected in 67.39% of the isolates (93/138). Three gene cassette arrays and 5 antimicrobial resistance gene cassettes were detected in 69 of the class 1 integron-positive strains. The most common gene cassette array was dfrA17-aadA5. Of the 93 intI1-positive strains, 5 different common promoters were detected. The most prevalent common promoter was PcH1, and most isolates contained the dfrA17-aadA5 gene cassette array. In summary, antimicrobial resistance and MDR were prevalent among E. coli isolates in our city Weifang in Shandong Provence China. Gene cassettes of the class 1 integron variable region mostly conferred resistance to traditional antimicrobials. Weak promoters in the variable regions were predominant in this study. Integrons pose a great threat to the treatment of MDR bacterial infections and further investigations are needed.

Distribution of integrons and antimicrobial resistance in carbapenem-resistant Acinetobacter baumannii clinical strains

Objective To investigate the distribution of integrons in clinical isolates of carbapenem-resistant Acinetobacter baumannii and their relationships to bacterial resistance to antimicrobial agents. Methods A total of 115 carbapenem-resistant Acinetobacter baumannii strains were isolated from clinical samples of patients from January to October, 2017. Phoenix 100 automatic microbiological analyzer was used for antimicrobial sensitivity analysis. Classes 1 and 2 integrase genes and carbapenemase-encoding genes, blaIMP, blaVIM, blaKPC, blaNDM and blaOXA-23, were screened by PCR. The variable regions of integrons were amplified by long fragment PCR. The types of promoters and gene cassette arrays of variable regions were determined by sequencing and overlap PCR. Relationships between integrons and antimicrobial resistance were analyzed. Results The 115 isolates of carbapenem-resistant Acinetobacter baumannii were resistant to most commonly used antimicrobial agents, but sensitive to polymyxin E. All of the isolates carried blaOXA-23 gene and none of them were positive for blaIMP, blaVIM, blaKPC or blaNDM gene. Class 1 integrase gene intI1 was detected in 40 isolates (34.8%), while class 2 integrase gene intI2 was not detected. Two gene cassette arrays of variable regions, aacA4-catB8-aadA1 (39 isolates) and aacC1-gacP-gacQ-aadA1a (23 isolates), were detected in intI1-positive isolates. Twenty-two isolates carried both aacA4-catB8-aadA1 and aacC1-gacP-gacQ-aadA1a. The upstream promoters of the variable regions were relatively strong promoters, PcH2 and PcS. The gene cassettes of the variable regions endowed bacteria with resistance to chloramphenicol and aminoglycoside antibiotics. The resistance rate of class 1 integron-positive isolates to compound sulfamethoxazole was higher than that of negative strains. However, their resistance rate to ampicillin/sulbactam was lower than that of negative strains. Conclusions Antimicrobial resistance in carbapenem-resistant Acinetobacter baumannii was serious. Carbapenem resistance was associated with blaOXA-23 gene. The types of promoters of variable regions in class 1 integrons were all relatively strong promoters. Class 1 integrons were closely related to sulfonamides resistance. Key words: Acinetobacter baumannii; Carbapenem; Antimicrobial resistance; Integron; Promoter

Characterization of Integrons and qnr Genes in Proteeae from a Teaching Hospital in China

Background: Proteeae isolates displaying multidrug-resistance (MDR) are the second most common causes of hospital-associated infections. The aim of this study was to screen class 1-3 integrons and plasmid-mediated quinolone resistance (PMQR) genes in Proteeae isolates from the First Affiliated Hospital of the Wenzhou Medical University. Materials and Methods: 176 Proteeae isolates were collected from clinical specimens of inpatients between January 2011 and December 2013. Susceptibility testing was determined by the agar dilution method. Class 1-3 integrons and PMQR genes were amplified by polymerase chain reaction, and the variable regions of integrons were determined by restriction fragment length polymorphisms. Results: 68.2% Proteeae isolates exhibited MDR phenotypes: 46.6 and 10.8% Proteeae isolates were positive for intI1 and intI2, respectively. The resistance rate of integron-positive isolates to aminoglycosides, fluoroquinolones, and trimethoprim/sulfamethoxazole was significantly higher than integron-negative isolates. Sequence analysis revealed that dfrA1-sat2-aadA1, dfrA1-catB2-sat2-aadA1, and sat2-aadA1 were first detected in Morganella morganii strains isolated from China. PMQR was determined by qnrD in 40 strains (22.7%). Conclusion: Our results indicate that class 1 and 2 integrons are common among Proteeae isolates. Meanwhile, qnrD are highly prevalent in Proteeae isolated from our hospital.

Multidrug resistance of enteric bacilli and its relation to structure and molecular evolution of variable region in resistance-related class-I integron

To investigate the drug resistance of enteric bacilli and its relation to the drug resistance gene cassette in the variable region and molecular evolution of class-I integron. K-B assay was applied to measure the drug resistance of E.coli, E.cloacae and A.baumannii isolated against twelve antibiotics. The class-I integron and drug resistance gene cassettes in the variable region of the integron were detected by PCR and sequencing of amplification products. The molecular evolution of drug resistance genes in the class-I integrons was analyzed using Clustal X and MEGA software. 54.2%-100% of A.baumannii isolates were resistant to the penicillin and cephem antibiotics, while E.coli and E.cloacae isolates had resistance rates of 41.6%-62.5% to cephem antibiotics. 62.5%(15/24) of E.coli, 67.9%(19/28) of E.cloacae and 83.3%(20/24) of A.baumannii isolates were positive for class-I integrons. 81.5% (44/54) of class-I integrons showed 4 different single band spectrums and the other class-I integrons displayed 3 different double band spectrums. In the drug resistance gene cassettes in variable regions of class-I integrons there were 7 types in 4 groups of drug resistance genes, including aac(6'), sad(3"), aad(2"), cat(4') and dfr (types 7, A13 and 15), which induced the resistance to aminoglycosides and sulfamido antibiotics and chloromycin. The class-I integrons in the isolates might be divided into 4 molecular evolution groups according to the diversity of dihydrofolate reductase encoding gene sequences. The enteric bacilli have a high drug resistance and frequently carry class-I integrons with 7 drug resistance gene cassettes which present 4 different evolutionary pathways.

Analysis of Antibiotic Resistance in Bacteria Isolated from the Surface Microlayer and Underlying Water of an Estuarine Environment

We compared the prevalence of cultivable antibiotic-resistant bacteria and resistance genes in the surface microlayer (SML) and underlying waters (UW) of an estuary. Prevalence of resistant bacteria was determined in antibiotic-supplemented agar. Bacterial isolates from the UW (n=91) and SML (n=80), selected in media without antibiotic, were characterized concerning susceptibility against nine antibiotics. The presence of genes bla(TEM), bla(OXA-B), bla(SHV), bla(IMP), tet(A), tet(B), tet(E), tet(M), cat, sul1, sul2, sul3, aadA, IntI1, IntI2, and IntI3 was assessed by PCR. The variable regions of integrons were sequenced. Ampicillin- and streptomycin-resistant bacteria were significantly more prevalent in SML. Resistance levels among the bacterial collections were generally low, preventing detection of significant differences between SML and UW. The tet(E) gene was detected in two Aeromonas isolates and tet(M) was detected in a Pseudomonas isolate. Gene sul1 was amplified from three Aeromonas isolates. Prevalence of intI genes was 2.11%. Cassette arrays contained genes encoding resistance to aminoglycosides and chloramphenicol. A higher prevalence of antibiotic-resistant bacteria in the SML, although only detectable when bacteria were selected in antibiotic-supplemented agar, suggests that SML conditions select for antibiotic resistance. Results also showed that antibiotic resistance was uncommon among estuarine bacteria and the resistance mechanisms are probably predominantly intrinsic.

Comment on: AbaR4 replaces AbaR3 in a carbapenem-resistant Acinetobacter baumannii isolate belonging to global clone 1 from an Australian hospital

Sir, I read with great interest the valuable article by Hamidian and Hall regarding the AbaR4 genomic island in D36, an Acinetobacter baumannii isolate belonging to global clone (GC) 1, and proposing a new family of transposons. However, I would like to make some remarks on the authors’ statement that D36 was of a rare sequence type (ST), ST247, and represented a new GC 1 lineage. In their study, the ST of A. baumannii strain D36 was determined by the Oxford multilocus sequence typing (MLST) scheme, which employs gltA, gyrB, gdhB, recA, cpn6, gpi and rpoD as housekeeping genes. As a result, strain D36 showed the allelic profile 10-12-4-11-4-58-5 and was assigned to ST247, slightly different from ST109 (10-12-4-11-4-9-5), which includes the GC 1 reference strain RUH875 (A297). Therefore, the difference between ST247 and ST109 was in their gpi alleles. However, in a recent study it was demonstrated that the gene gpi resided in a part of the A. baumannii genome prone to recombination and showed significant variations compared with other housekeeping genes, and a similar observation was also described in another study. Thus it appears to be problematic to include gpi in the MLST system and therefore the conclusion that ST109 and ST247 are two distinct lineages is also doubtful. Besides AbaR4-associated features, such as lack of Tn6018 and the resistance genes catA1, blaTEM, tet(A) and aphA1, strain D36 also showed other characteristics (aadB gene cassette and sul2 gene) different from other GC 1 strains included in the study by Hamidian and Hall. It has been suggested that the accessory genome (genes present in some isolates, but not in others) can be used as a tool for epidemiological investigation, and that the distinct features of strain D36 might support its role as the representative of a new lineage. On the one hand, it is difficult to select epidemiological markers from the hundreds of genes in the accessory genome, which varies between isolates. On the other hand, comparative genomic analysis has demonstrated that there are frequent homologous recombination events across up to 20% of GC 2 genomes, and the recombination also took place across a large phylogenetic distance, including GC 1. The horizontal transfer of aminoglycoside resistance genes and gene cassettes representing the variable region of integrons between GC 1 and 2 isolates from the Czech Republic has also been described. Therefore, the usefulness of the accessory genome in epidemiological research remains to be determined. As a result, the taxonomic position of strain D36 should be determined by a new scheme, for example, the MStree analysis-based MLST system, and further evidence is required to support the conclusion that strain D36 belongs to a new GC 1 lineage.

Effect of attC structure on cassette excision by integron integrases

BackgroundIntegrons are genetic elements able to integrate and disseminate genes as cassettes by a site-specific recombination mechanism. These elements contain a gene coding for an integrase that carries out recombination by interacting with two different target sites; the attI site in cis with the integrase and the palindromic attC site of a gene cassette. Integron integrases (IntIs) bind specifically to the bottom strand of attC sites. The extrahelical bases resulting from folding of attC bottom strands are important for the recognition by integrases. These enzymes are directly involved in the accumulation and formation of new cassette arrangements in the variable region of integrons. Thus, it is important to better understand interactions between IntIs and their substrates.ResultsWe compared the ability of five IntIs to carry out excision of several cassettes flanked by different attC sites. The results showed that for most cassettes, IntI1 was the most active integrase. However, IntI2*179E and SonIntIA could easily excise cassettes containing the attCdfrA1 site located upstream, whereas IntI1 and IntI3 had only a weak excision activity for these cassettes. Analysis of the secondary structure adopted by the bottom strand of attCdfrA1 has shown that the identity of the extrahelical bases and the distance between them (A-N7-8-C) differ from those of attCs contained in the cassettes most easily excisable by IntI1 (T-N6-G). We used the attCdfrA1 site upstream of the sat2 gene cassette as a template and varied the identity and spacing between the extrahelical bases in order to determine how these modifications influence the ability of IntI1, IntI2*179E, IntI3 and SonIntIA to excise cassettes. Our results show that IntI1 is more efficient in cassette excision using T-N6-G or T-N6-C attCs while IntI3 recognizes only a limited range of attCs. IntI2*179E and SonIntIA are more tolerant of changes to the identity and spacing of extrahelical bases.ConclusionsThis study provides new insights into the factors that influence the efficiency of cassette excision by integron integrases. It also suggests that IntI2 and SonIntIA have an evolutionary path that is different from IntI1 and IntI3, in their ability to recognize and excise cassettes.

Reservoir of Antimicrobial Resistance Determinants Associated with Horizontal Gene Transfer in Clinical Isolates of the Genus Shewanella

Although Shewanella is usually considered an environmental genus, different clinical infections have appeared in recent years (4, 7). Treatment of such infections is difficult due to the lack of knowledge concerning the natural antimicrobial resistance as well as the recommended antibiotic treatment of their infections (11). The aim of our study was to investigate the antimicrobial resistance mechanisms acquired by this genus in the nosocomial environment. All isolates identified as Shewanella spp. (n = 10) by the use of standard biochemical tests were collected in a public hospital of Argentina during 2005 and 2006. In Argentina, the relative frequency of isolates positively identified as harboring Shewanella spp. among infections by nonfermentative Gram-negative bacilli (NFGNB), excluding the important pathogens Acinetobacter baumannii and Pseudomonas aeruginosa, is only 2.5% of the total number of isolates. Amplification and sequencing of the 16S rRNA gene was used to identify the isolates (4, 12). Three isolates were identified as Shewanella putrefaciens and seven as Shewanella algae (Table ​(Table11). TABLE 1. Antimicrobial resistance mechanisms found among Shewanella clinical isolatesa PCR amplifications using total DNA were performed according to the instructions of the manufacturer (Promega) by the use of specific primers for evaluation of the presence of antimicrobial resistance determinants associated with horizontal gene transfer (most of them usually found in our Gram-negative bacterial isolates): (i) integron integrase genes (intI1, intI2, and intI3) (6, 10); (ii) 13 β-lactamase-like genes, namely, the blaOXA-58, blaOXA-48, blaTEM-1, blaCTX-M-2, blaSHV-like, blaSCO-1, blaPER-2, blaGES-like, blaVEB-like, blaIMP-like, blaVIM-like, blaSPM-1, and blaOXA-23 genes (5, 6, 9); and (iii) sul1, sul2, and sul3 genes (1). We found the type 1 integrase gene in 9 isolates, while the type 2 integrase gene was found in 6 isolates (Table ​(Table1).1). In order to identify the inserted gene cassettes within the variable region of integrons, PCR cartography and sequencing were performed as previously described (6, 10). Concerning the variable region of class 1 integrons (vr-1), we found the presence of the dfrA1 gene cassette in isolates Sa9, Sa82, and Sa392 and the dfrA1-aadA1 array in isolates Sa74, Sa10, and Sa2 (Table ​(Table1).1). We were unable to identify gene cassettes within the vr-1 for three isolates (Sp117, Sp95, and Sp31). Concerning class 2 integrons, only the variable region of Sa2 harboring dfrA1-sat2-aadA1-orfX-ybfA-ybfB-ybgA, Sa9 harboring dfrA1-sat2, and Sa10 harboring the dfrA1 gene cassette could be identified (Table ​(Table11). When we investigated the presence of 13 β-lactamase genes, positive amplification was obtained for the 3 isolates of S. putrefaciens (Table ​(Table1)1) by the use of primers for the amplification of the carbapenemase blaOXA-48 gene previously found harbored in a transposon of a Klebsiella pneumoniae isolate from France (9). Sequence analysis revealed 99% identity over a 690-bp length with this gene, 83% identity with the sequence of blaOXA-54 previously described as present in S. oneidensis and suggested as the progenitor of the blaOXA-48 (8), and 78% identity with the sequence of a chromosome-borne blaOXA gene that we have identified in S. putrefaciens CN-32 ({type:entrez-nucleotide,attrs:{text:CP000681.1,term_id:145562801,term_text:CP000681.1}}CP000681.1) by the use of BLAST software (version 2.0). The meropenem (MEM) and imipenem (IMP) MICs were determined according to CLSI guidelines (3). As was shown in Table ​Table1,1, no clear contribution of this gene to carbapenem resistances could be established. Since there is no information about the occurrence of plasmids in Shewanella spp., we performed plasmid extraction using a QIAprep Spin Miniprep kit (Qiagen) to see whether our isolates contained plasmids. We found that plasmids were present in 2 out of 10 isolates (Sa2 and Sp95). We performed PCRs for commonly incompatible groups (IncP, IncW, and IncA/C) found in our bacterial population (reference 2 and data not shown) as well as plasmid extraction followed by Escherichia coli transformation and selection with the corresponding antibiotics, but all experiments gave negative results. We found not only high levels of dispersion of genetic elements usually associated with horizontal gene transfer for both species but also distinctive epidemiology characteristics, since S. putrefasciens isolates possess the blaOXA-48 gene and class 1 integrons, while S. algae isolates possess class 1 and 2 integrons with different arrays of cassettes in the two species (Table ​(Table11). Given that Shewanella is well known as an environmental genus and that almost all isolates from our study harbored integrons and other relevant determinants of resistance (such as the carbapenemase blaOXA-48) usually associated with horizontal gene transfer, species of this genus could be considered to represent not only a potential reservoir but also a vector of antimicrobial resistance mechanisms in hospital settings and the environment, with transmission occurring in both directions.

Characterization of Salmonella enterica serovar Typhimurium conjugative plasmids transferring resistance to antibiotics and their interaction with the virulence plasmid

In this study, we analysed field isolates of Salmonella enterica serovar Typhimurium for the presence of conjugative plasmids transferring resistances to antibiotics. Altogether 23 strains were analysed for the presence of conjugative R-plasmids. In the case of successful conjugation, the R-plasmids were characterized by PCR for antibiotic resistance genes, integrons and replicon typing. Variable regions of integrons were sequenced. Conjugation and transfer of antibiotic resistance was observed in 12 strains. Conjugative plasmids in these strains belonged to the IncI1 and IncHI1 replicons and four of them transferred antibiotic resistance associated with class I integrons. In two cases, resistance to tetracycline and/or ampicillin was not transferred by conjugation to approximately 10% of the transconjugants. Detailed characterization showed that the loss of both resistances was associated with the loss of Tn3 (bla(TEM)) and Tn1721 [tet(A)] from the conjugative plasmids p9046 and p9134. However, when only the tetracycline resistance was lost, the Tn1721 was replaced with a partial sequence of rck, and with complete coding sequences of srgA, srgB, ORF7 and pefI originating from the Salmonella Typhimurium virulence plasmid. Two plasmids from our collection were capable of recombination with the virulence plasmid of Salmonella Typhimurium and subsequently spread both antibiotic resistance and virulence genes to the recipient.

Prevalence of aadA1 and dfrA15 class 1 integron cassettes and SXT circulation in Vibrio cholerae O1 isolates from Africa

Prevalence of aadA1 and dfrA15 class 1 integron cassettes and SXT circulation in Vibrio cholerae O1 isolates from Africa

Study on the mechanism of integron mediated multi-resistance in E. coli and Klebsiella

目的 研究整合子参与大肠埃希菌和克雷伯菌多重耐药的分子机制.方法 纸片扩散法测定61株临床分离的大肠埃希菌和克雷伯菌的药物敏感性;整合酶基因扩增法检测Ⅰ类、Ⅱ类和Ⅲ类整合子;整合子可变区扩增并测序.结果 73.8%(45/61)的菌株Ⅰ类整合子阳性,2株大肠埃希菌Ⅱ类整合子阳性,未检测到Ⅲ类整合子;88.9%(40/45)的Ⅰ类整合子阳性菌株可变区扩增阳性,扩增片段大小从150～2 800 bp不等,有1株Ⅱ类整合子阳性菌株可变区扩增阳性,大小为2 200bp;整合子可变区含有编码对氨基糖苷类抗生素耐药的基因(aadA1、aadA2、aadA5、aadB、aacA4、aac6'-Ib)、编码对磺胺类抗生素耐药的基因(dfrA1、dfr2d、dfrⅤ、dfrⅫ、dfr17)、编码对氯霉素耐药的基因(cat、catB8、cmlA1-variant)、编码对β-内酰胺类抗生素耐药的基因(blaoxa10)和编码对链丝菌素耐药的基因(sat1).结论 整合子在大肠埃希菌和克雷伯菌中广泛存在,参与了这两种菌多重耐药的形成。

Characterization of multidrug-resistance phenotypes and genotypes of Escherichia coli strains isolated from swine from an abattoir in Osaka, Japan

A total of 455 highly tetracycline-resistant Escherichia coli strains were isolated from 84 healthy swine from abattoirs and it was found that 56.9, 43.1, 22.2, 15.4, 2.6 and 1.5% of strains were resistant to chloramphenicol, ampicillin, kanamycin, trimethoprim-sulphamethoxazole, ofloxacin and gentamicin respectively. Interestingly, E. coli strains isolated from certain finisher hog groups exhibited resistance against 2-7 antimicrobials, but strains isolated from multiparous sow groups in each herd were resistant to only 2-4 antimicrobial agents. When randomly selected 108 tetracycline-resistant isolates were tested for the presence of resistance genes, the following genes tet(A) (n = 6), tet(B) (n = 95), tet(D) (n = 1) or both tet(A) and tet(B) (n = 6) were found to be distributed among them. Furthermore, 52 isolates carried the integrase 1 gene and 24 strains gave five different PCR amplicon profiles using primers from the variable region of integron. Extensive nucleotide sequence analyses of these amplicons revealed the presence of dhfrI, dhfrXII, dfr17, aadA, aadA2, aadA5, aadA21, aacA4 and catB3 genes which code for different antibacterial resistance proteins.

Site-specific deletion and rearrangement of integron insert genes catalyzed by the integron DNA integrase.

Deletion of individual antibiotic resistance genes found within the variable region of integrons is demonstrated. Evidence for gene duplications and rearrangements resulting from the insertion of gene units at new locations is also presented. Deletion, duplication, and rearrangement occur only in the presence of the integron-encoded DNA integrase. These events are precise and involve loss or gain of one or more complete insert units or gene cassettes. This confirms the recent definition of gene cassettes as consisting of the gene coding sequences, all except the last 7 bases of the 59-base element found at the 3' end of the gene, and the core site located 5' to the gene (Hall et al., Mol. Microbiol. 5:1941-1959, 1991) and demonstrates that individual gene cassettes are functional units which can be independently mobilized. Both deletions and duplications can be generated by integrase-mediated cointegrate formation followed by integrase-mediated resolution involving a different pair of sites. However, deletion occurs 10 times more frequently than duplication, and we propose that the majority of deletion events are likely to involve integrase-dependent excision of the gene unit to generate a circular gene cassette. The implications of these findings in understanding the evolution of integrons and the spread of antibiotic resistance genes in bacterial populations is discussed.