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PubMed1994

Integrons found in different locations have identical 5' ends but variable 3' ends.

The positions of the outer boundaries of the 5'- and 3'-conserved segment sequences of integrons found at several different locations have been determined. The position of the 5' end of the 5'-conserved segment is the same for six independently located integrons, In1 (R46), In2 (Tn21), In3 (R388), In4 (Tn1696), In5 (pSCH884), and In0 (pVS1). However, the extent of the 3'-conserved segment differs in each integron. The sequences of In2 and In0 diverge first from the conserved sequence, and their divergence point corresponds to the 3'-conserved segment endpoint defined previously (H.W. Stokes and R.M. Hall, Mol. Microbiol. 3:1669-1683, 1989), which now represents the endpoint of a 359-base deletion in In0 and In2. The sequence identity in In3, In1, In4, and In5 extends beyond this point, but each sequence diverges from the conserved sequence at a different point within a short region. Insertions of IS6100 were identified adjacent to the end of the conserved region in In1 and 123 bases beyond the divergence point of In4. These 123 bases are identical to the sequence found at the mer end of the 11.2-kb insertion in Tn21 but are inverted. In5 and In0 are bounded by the same 25-base inverted repeat that bounds the 11.2-kb insert in Tn21, and this insert now corresponds to In2. However, while In0, In2, and In5 have features characteristic of transposable elements, differences in the structures of these three integrons and the absence of evidence of mobility currently preclude the identification of all of the sequences associated with a functional transposon of this type.

Base Sequence↗
PubMed1996

The integrons In0, In2, and In5 are defective transposon derivatives.

The class 1 integrons In0, In2, and In5, found in different locations in pVS1, Tn21, and pSCH884, have closely related structures. All three integrons contain an insertion sequence, IS1326, that is a new member of the IS21 family. IS1326 has caused deletions of adjacent 3'-conserved segment and transposition module sequences, and all three integrons retain a complete copy of only one of four genes required for transposition of related transposons and are thus defective transposon derivatives. In2 contains an additional insertion sequence, IS1353, located within IS1326. IS1353 is a member of the IS3 family and appears to have been acquired after the integron was inserted into an ancestral mercury resistance transposon to create the ancestor of Tn21 and several other transposons that are close relatives of Tn21.

Amino Acid Sequence↗
PubMed2001

Characterization of In53, a class 1 plasmid- and composite transposon-located integron of Escherichia coli which carries an unusual array of gene cassettes.

Further characterization of the genetic environment of the gene encoding the Escherichia coli extended-spectrum beta-lactamase, bla(VEB-1), revealed the presence of a plasmid-located class 1 integron, In53, which carried eight functional resistance gene cassettes in addition to bla(VEB-1). While the aadB and the arr-2 gene cassettes were identical to those previously described, the remaining cassettes were novel: (i) a novel nonenzymatic chloramphenicol resistance gene of the cmlA family, (ii) a qac allele encoding a member of the small multidrug resistance family of proteins, (iii) a cassette, aacA1b/orfG, which encodes a novel 6'-N-acetyltransferase, and (iv) a fused gene cassette, oxa10/aadA1, which is made of two cassettes previously described as single cassettes. In addition, oxa10 and aadA1 genes were expressed from their own promoter sequence present upstream of the oxa10 cassette. arr-2 coded for a protein that shared 54% amino acid identity with the rifampin ADP-ribosylating transferase encoded by the arr-1 gene from Mycobacterium smegmatis DSM43756. While in M. smegmatis, the main inactivated compound was 23-ribosyl-rifampin, the inactivated antibiotic recovered from E. coli culture was 23-O-ADP-ribosyl-rifampin. The integrase gene of In53 was interrupted by an IS26 insertion sequence, which was also present in the 3' conserved segment. Thus, In53 is a truncated integron located on a composite transposon, named Tn2000, bounded by two IS26 elements in opposite orientations. Target site duplication at both ends of the transposon indicated that the integron likely was inserted into the plasmid through a transpositional process. This is the first description of an integron located on a composite transposon.

Amino Acid Sequence↗
PubMed2002

The IntI-like tyrosine recombinase of Shewanella oneidensis is active as an integron integrase.

We have found an integron-like integrase gene and an attI site in Shewanella oneidensis as part of a small chromosomal integron. We have cloned this gene and tested the ability of the integrase to excise cassettes from various integrons. Most cassettes flanked by two attC sites are readily excised, while cassettes in the "first" position, with an attI1 or attI3 site on one end, are not excised. An exception is a cassette with attI2 on one end. The attI2 site, from Tn7, has greater similarity to the attI site adjacent to the integrase of S. oneidensis than do attI1 or attI3. We cloned the attI site of S. oneidensis and observed the integration of two different cassettes. We have, therefore, demonstrated the function of this integron-like integrase.

Amino Acid Sequence↗
PubMed2001

Molecular epidemiology of the integron-located VEB-1 extended-spectrum beta-lactamase in nosocomial enterobacterial isolates in Bangkok, Thailand.

Over a 21/2-month period in 1999, 37 ceftazidime-resistant nonrepetitive enterobacterial isolates were collected from 37 patients in a Bangkok hospital, Thailand. Eighty-one percent of these strains expressed a clavulanic acid-inhibited extended-cephalosporin resistance profile. An identical extended-spectrum beta-lactamase (ESBL), VEB-1, was found in 16 unrelated enterobacterial isolates (Escherichia coli, n = 10; Enterobacter cloacae, n = 2; Enterobacter sakazakii, n = 1; and Klebsiella pneumoniae, n = 3) and in two clonally related E. cloacae isolates. The bla(VEB-1) gene was located on mostly self-conjugative plasmids (ca. 24 to 200 kb) that conferred additional non-beta-lactam antibiotic resistance patterns. Additionally, the bla(VEB-1) gene cassette was part of class 1 integrons varying in size and structure. The bla(VEB-1)-containing integrons were mostly associated with bla(OXA-10)-like and arr-2-like gene cassettes, the latter conferring resistance to rifampin. These data indicated the spread of bla(VEB-1) in Bangkok due to frequent transfer of different plasmids and class 1 integrons and rarely to clonally related strains. Plasmid- and integron-mediated resistance to rifampin was also found in enterobacterial isolates.

Anti-Bacterial Agents↗
PubMedDate not supplied

Rapid emergence of ciprofloxacin-resistant enterobacteriaceae containing multiple gentamicin resistance-associated integrons in a Dutch hospital.

In a hematology unit in the Netherlands, the incidence of ciprofloxacin-resistant Enterobacter cloacae and Escherichia coli increased from from 1996 to 1999. Clonal spread of single genotypes of both ciprofloxacin-resistant E. coli and Enterobacter cloacae from patient to patient was documented by pulsed-field gel electrophoresis and random amplification of polymorphic DNA. In addition, genetically heterogeneous strains were isolated regularly. Integrons associated with gentamicin resistance were detected in Enterobacter cloacae and E. coli strains. Integron-containing E. coli were detected in all hematology wards. In contrast, in Enterobacter cloacae strains two integron types were encountered only in the isolates from one ward. Although in all patients identical antibiotic regimens were used for selective decontamination, we documented clear differences with respect to the nosocomial emergence of ciprofloxacin-resistant bacterial strains and gentamicin resistance-associated integrons.

Anti-Bacterial Agents↗
PubMed2004

Molecular characterization of the integrons in Shigella strains isolated from patients with traveler's diarrhea.

The prevalence and characterization of Class 1 integrons has been performed in eighty three strains of Shigella spp., isolated between 1995 and 2000 from patients with traveler's diarrhea. A low prevalence (13.25%) was recorded. Nine different integrons were found among 11 multiresistant strains, with a total of 10 different gene cassettes encoding for resistance to trimethoprim (dfrA1, dfrA5, dfrA7, dfrA12, and dfrA15), aminoglycosides (aadA1a and aadA2), beta-lactam antibiotics (oxa2) or ORF with unknown function (orfD and orfF). A high prevalence of dfr and aad gene cassettes was observed. The low incidence of Class 1 integrons observed in this study is in contrast with the known facility that the Shigella genus has to gain and transfer plasmids.

Anti-Bacterial Agents↗
PubMed2005

Characterization of class 1 integrons-mediated antibiotic resistance among calf pathogenic Escherichia coli.

Escherichia coli isolates from calf diarrhea cases (n=22) in the Beijing surrounding region in China were characterized for disease serotype, virulence factors, antimicrobial susceptibility pattern and class 1 integrons. 59% (n=13) of the isolates were positive for the int I1 gene. The presence and genetic content of class 1 integrons in 13 E. coli isolates were examined by PCR and sequencing. Sequencing analysis revealed six gene cassettes, which encoded resistance to trimethoprim (dfrA1, dfrA17), aminoglycosides (aadB, aadA1 and aadA5) and chloramphenicol (cmlA). The gene cassette arrays dfrA1-orf (45%) and aadB-orf-cmlA (32%) were most prevalent among these isolates. These data revealed the high prevalence of class 1 integrons among calf pathogenic E. coli isolates in the Beijing surrounding region in China, which may provide important and useful surveillance information reflecting specific antibiotic selective pressure.

Animals↗
PubMed2004

Detection of integrons and antibiotic-resistance genes in Salmonella enterica serovar Typhimurium isolates with resistance to ampicillin and variable susceptibility to amoxicillin-clavulanate.

We characterized 29 antimicrobial-resistant Salmonella enterica serovar Typhimurium strains, including four belonging to the monophasic variant 4,5,12:i:-, mostly isolated from infants. They were selected from 3230 strains isolated in the years 1990-2001 on the basis of resistance to ampicillin and variable susceptibility to the amoxicillin-clavulanate combination. Twenty-three strains were resistant to more than four antibiotics. All the strains carried the bla(TEM) gene and most were able to transfer this gene by conjugation. Sequencing of the gene from one of the amoxicillin-clavulanate-resistant strains allowed identification of the encoded beta-lactamase as TEM-1; all of these strains carried a second gene encoding beta-lactamase production, either pse-1 or oxa1. However, the association of bla(TEM) plus pse-1 genes did not always confer resistance to amoxicillin-clavulanate. The pse-1 gene, found in 17 strains, was located in the Salmonella Genomic Island-1 (SGI1), which carries two integrons and encodes multiple drug-resistance. None of the oxa1-bearing strains had the SGI1, yet this gene was found as part of an integron that also carried the aadA1 gene and was not plasmid-associated. Thirteen of the strains harbouring SGI1 belonged to the definitive phage type (DT) 104, and most of those remaining to DT104b and U302; particularly, strains carrying the oxa1-aadA1 integron belonged to the last two phage types. Pulsed field electrophoresis confirmed the clonal organization of DT104 strains, whereas U302 strains fell into different groups, depending on their resistance determinants.

Amoxicillin↗
PubMed2006

Integron presence in a multiresistant Morganella morganii isolate.

A multiresistant strain of Morganella morganii was isolated from a patient affected by several severe pathologies. The isolate was found to be resistant to the following antimicrobials: ampicillin, nalidixic acid, cefalothin, cefoxitin, ceftriaxone, ciprofloxacin, chloramphenicol, streptomycin, erythromycin, gentamicin, novobiocin, penicillin, rifampicin, tetracycline and violet crystal. Mechanisms leading to this multiresistance were studied. Porins of M. morganii multiresistant and wild-type strains were analysed by sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and were characterised by their ability to form channels in planar black lipid bilayers. The channels formed by porins from multiresistant and susceptible strains suggested that the porins of the multiresistant strain were not responsible for resistance. A 6.6 kb plasmid (pML2003) was detected, isolated and studied. pML2003 included two integrons. Direct sequencing revealed that one of the integrons contained two cassettes, aminoglycoside adenyltransferase (aadB) and chloramphenicol acetyltransferase (catB3) conferring resistance to aminoglycosides and chloramphenicol, respectively. The second integron contained carbenicillinase (blaP1b) and adenyltransferase (aadA2), which confer resistance to beta-lactamases and streptomycin, respectively.

Drug Resistance, Multiple, Bacterial↗
PubMed2005

Multiplex PCR for screening of integrons in bacterial lysates.

Bacterial integrons are a useful PCR amplification target in epidemiological surveys of bacterial antibiotic resistance, and a variety of primers have been published. We describe multiplex PCR methodology to test for classes 1, 2 and 3 integron-associated integrases in boiled lysates of Gram-negative bacteria. We report on performance in Acinetobacter spp. (n=50), Enterobacteriaceae (n=76), Pseudomonas aeruginosa (n=15), Bacteroidesspp. (n=69), and in undifferentiated mixed cultures derived from perineal swabs (n=50) and endotracheal aspirates (n=8). This method achieved 100% sensitivity and specificity in simple lysates made from a range of bacteria, without requiring DNA extraction, and is recommended as an efficient screening tool for surveys of integron cassettes.

Acinetobacter↗
PubMed2004

Sequence of the 68,869 bp IncP-1alpha plasmid pTB11 from a waste-water treatment plant reveals a highly conserved backbone, a Tn402-like integron and other transposable elements.

To analyse the significance of conjugative broad-host-range IncP-1alpha plasmids for the spread of antibiotic resistance determinants in waste-water treatment plants we isolated and characterised five different IncP-1alpha plasmids from bacteria of activated sludge and the final effluents of a municipal waste-water treatment plant. These plasmids mediate resistance to ampicillin, cefaclor, cefuroxime, gentamicin, kanamycin, spectinomycin, streptomycin, tetracycline, tobramycin, and trimethoprim. The complete 68,869 bp DNA-sequence of the IncP-1alpha plasmid pTB11 was determined. The pTB11 backbone modules for replication (Rep), mating pair formation (Trb), multimer resolution (Mrs), post-segregational killing (Psk), conjugative DNA-transfer (Tra), plasmid control (Ctl), and stable maintenance and inheritance (KilA, KilE, and KilC) are highly conserved as compared to the 'Birmingham' IncP-1alpha plasmids. In contrast to the 'Birmingham' plasmids pTB11 carries an insert of a Tn402-derivative integrating a class 1 integron in the intergenic region between the multimer resolution operon parCBA and the post-segregational killing operon parDE. The integron comprises the resistance gene cassettes oxa2 (beta-lactamase), aacA4 (aminoglycoside-6'N-acetyltransferase), and aadA1 (aminoglycoside-3'-adenylyltransferase) and a complete tniABQR transposition module. Integron-specific sequences were also identified on other IncP-1alpha plasmids analysed in this work. In contrast to the 'Birmingham' plasmids the pTB11 tetracycline resistance module carries a pecM- and a pncA-like gene downstream of the tetracycline resistance gene tetA and contains an insertion of the new insertion sequence element ISTB11. The transposable elements IS21 and Tn1 which disrupted, respectively, orf7 and klcB on the 'Birmingham' plasmids are not present on pTB11. Identification of IncP-1alpha plasmids in bacteria of the waste-water treatment plant's final effluents indicates that bacteria carrying these kind of plasmids are released into the environment.

Base Sequence↗
PubMed2003

Characterization of class 1 integron resistance gene cassettes and the identification of a novel IS-like element in Acinetobacter baumannii.

Based on hybridization studies, 21/32 multi-resistant clinical isolates of Acinetobacter baumannii contain class 1 integrons. Amplification products were obtained from 20 of the hybridization-positive strains. A single dfrA7 cassette was identified in 18 of the isolates and an integron with two cassettes (aadB-aadA4) was found in only one strain. Amplicons were not obtained from one of the hybridization positive strains. DNA sequence analysis of a 6.080-kb fragment, cloned from this strain, identified the remnant of an integron, following insertion of IS26 into the 5(')-end of intI1. The 6.080-kb sequence carries an aminoglycoside resistance gene, linked to a portion of IS1133, which in turn is linked to a sequence that has properties of IS elements, including sequences that could stimulate transcription, and ORFs encoding amino acid sequences with similarity to a transposase from Deinococcus radiodurans.

Acinetobacter baumannii↗
PubMed2006

Integrons: agents of bacterial evolution.

Integrons are assembly platforms - DNA elements that acquire open reading frames embedded in exogenous gene cassettes and convert them to functional genes by ensuring their correct expression. They were first identified by virtue of their important role in the spread of antibiotic-resistance genes. More recently, our understanding of their importance in bacterial genome evolution has broadened with the discovery of larger integron structures, termed superintegrons. These DNA elements contain hundreds of accessory genes and constitute a significant fraction of the genomes of many bacterial species. Here, the basic biology of integrons and superintegrons, their evolutionary history and the evidence for the existence of a novel recombination pathway is reviewed.

Attachment Sites, Microbiological↗
PubMed2004

Gram-positive bacteria are a major reservoir of Class 1 antibiotic resistance integrons in poultry litter.

Reversing the spread of antibiotic multiresistant bacteria is hampered by ignorance of the natural history of resistance genes, the mobile elements carrying them, and the bacterial hosts harboring them. Using traditional cultivation and cultivation-independent molecular techniques, we quantified antibiotic resistance genes and mobile elements called integrons in poultry house litter from commercial poultry farms. Unexpectedly, the major reservoir for Class 1 integrons in poultry litter is not their previously identified hosts, Gram-negative Enterobacteriaceae such as Escherichia coli. Rather, integrons and associated resistance genes abound in several genera of Gram-positive bacteria that constitute >85% of the litter community compared with Enterobacteriaceae that comprise <2% of this ecosystem. This finding warrants reexamination of our assumptions about the persistence and spread of antibiotic resistance genes.

Animals↗
PubMed2002

The role of integrons in antibiotic resistance gene capture.

Although recently discovered, integrons have played a primordial role in the evolution of bacterial genomes. They are best known as the genetic agents responsible for the capture and spread of antibiotic resistance determinants among diverse Gram-negative clinical isolates, and this activity is at the root of the antibiotic resistance phenomenon that has evolved over the last 60 years. The discovery of the ancestral chromosomal super-integrons, novel integron classes, and the multitude of gene cassettes they propagate solidify the crucial role of this system in adaptive bacterial evolution. Recent evidence suggests that evolutionarily old genetic recombination mechanisms for gene transfer have been adapted to the new antibiotic environment due to the heavy selective pressure of liberal antibiotic use in human medicine and animal husbandry.

Anti-Bacterial Agents↗
PubMed2003

New dfr2 gene as a single-gene cassette in a class 1 integron from a trimethoprim-resistant Escherichia coli isolate.

Resistance to trimethoprim among Enterobacteriaceae is increasing in spite of a stable or decreasing use of the drug. Integrons are common among these bacteria and many of them contain dfr gene cassettes. A clinical isolate of Escherichia coli from a urine specimen obtained at the Karolinska Hospital in Stockholm was resistant to trimethoprim, ampicillin, sulfonamides, chloramphenicol, streptomycin, and norfloxacin. PCR analysis with primers specific for the 5' and 3' sequences flanking the cassettes in class 1 integrons, detected a very short cassette region of no more than approximately 400 bp. Sequence analysis of the entire integron was performed, revealing a single-gene cassette of 411 bp encoding a new dihydrofolate reductase. The gene cassette comprised all sequences between the flanking conserved sequences and encoded only 78 amino acids. By homology it belongs to the unique group of dfr2 gene cassettes and being the fourth described gene in this group the new gene is called dfr2d. The enzymes encoded by the dfr2 genes are 67% identical and are not related to any other known dfr gene products.

Amino Acid Sequence↗
PubMed2004

Characterization and localization of drug resistance determinants in multidrug-resistant, integron-carrying Salmonella enterica serotype Typhimurium strains.

The genetic background of the antimicrobial resistance of 10 selected multiresistant Salmonella serotype Typhimurium (S. Typhimurium) strains (including the emerging monophasic variant [4,5,12:i:- ]) was investigated. All strains shared class 1 integrons (with seven types of variable regions) and belonged to different lineages (L1-L6) according to their phage types, DNA polymorphisms by XbaI-pulsed-field gel electrophoresis (PFGE), integrons, and/or resistance patterns. The strains were screened for the presence and localization (chromosomal or plasmid) of 32 DNA sequences representing integron-, Tn21-like transposon-, resistance-, and virulence-plasmid genes. Strains belonging to lineage L1 (definitive phage type DT104) carried the 90-kb Salmonella virulence plasmid together with the complete or partial chromosomally located Salmonella Genomic Island 1 (SGI1). All strains belonging to the other five lineages carried their resistance determinants on various resistance plasmids. Two of these strains showed complex plasmid profiles, which included a 95 kb virulence plasmid together with two or four resistance plasmids. Two strains carried a resistance plasmid that lacked the virulence-plasmid-encoding sequences. The remaining two strains carried two different hybrid virulence-resistance plasmids. Twenty-three of the DNA sequences could be assigned to distinct XbaI genomic restriction patterns (PFGE profiles). In this way, the influence of the resistance and virulence plasmids on the PFGE profiles was determined, and several groups of resistance genes could be identified. The data obtained represent a useful epidemiological tool for tracing the emergence and distribution of multiresistant S. Typhimurium worldwide.

Base Sequence↗