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IS1R-mediated plasticity of IncL/M plasmids leads to the insertion of bla OXA-48 into the Escherichia coli Chromosome.

ABSTRACT: The OXA-48 carbapenemase is mainly encoded by ? 62-kb IncL/M plasmids. However, chromosome-mediated genes have been observed in Escherichia coli isolates. In this work, we investigated the genetic environment of OXA-48 in members of the family Enterobacteriaceae (n = 22) to understand how the OXA-48-encoding gene is transferred into the E. coli chromosome. The OXA-48-encoding gene was located within intact Tn1999.2 transposons in the ? 62-kb plasmids or within a truncated variant of Tn1999.2 for the OXA-48-encoding genes located in the chromosomes of E. coli bacteria. The analysis of the Tn1999.2 genetic environment revealed an inverted orientation of the transposon in five ? 62-kb plasmids (5/14 [35%]) and in all chromosome inserts (n = 8). The sequencing of pRA35 plasmid showed that this orientation of Tn1999.2 and the acquisition of an IS1R insertion sequence generated a 21.9-kb IS1R-based composite transposon encoding OXA-48 and designated Tn6237. The sequencing of a chromosomal insert encoding OXA-48 also revealed this new transposon in the E. coli chromosome. PCR mapping showed the presence of this element in all strains harboring an OXA-48-encoding chromosomal insert. However, different insertion sites of this transposon were observed in the E. coli chromosome. Overall, these findings indicate a plasticity of the OXA-48 genetic environment mediated by IS1R insertion sequences. The insertion sequences can induce the transfer of the OXA-encoding gene into E. coli chromosomes and thereby promote its persistence and expression at low levels.

SUBMITTER: Beyrouthy R

PROVIDER: S-EPMC4068556 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature

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IS1R-mediated plasticity of IncL/M plasmids leads to the insertion of bla OXA-48 into the Escherichia coli Chromosome.

Beyrouthy R R Robin F F Delmas J J Gibold L L Dalmasso G G Dabboussi F F Hamzé M M Bonnet R R

Antimicrobial agents and chemotherapy 20140421 7

The OXA-48 carbapenemase is mainly encoded by ∼ 62-kb IncL/M plasmids. However, chromosome-mediated genes have been observed in Escherichia coli isolates. In this work, we investigated the genetic environment of OXA-48 in members of the family Enterobacteriaceae (n = 22) to understand how the OXA-48-encoding gene is transferred into the E. coli chromosome. The OXA-48-encoding gene was located within intact Tn1999.2 transposons in the ∼ 62-kb plasmids or within a truncated variant of Tn1999.2 for ...[more]

PMID: 24752261

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Project description:Introductionbla(OXA-48) is a globally emerging carbapenemase-encoding gene. The progenitor of bla(OXA-48) appears to be a Shewanella species. The presence of the bla(OXA-48)-like gene was investigated for two Shewanella xiamenensis strains.MethodsStrain WCJ25 was recovered from post-surgical abdominal drainages, while S4 was the type strain of S. xiamenensis. Species identification for WCJ25 was established by sequencing the 16S rDNA and gyrB genes. PCR was used to screen the bla(OXA-48)-like genes and to obtain their complete sequences. A phylogenetic tree of the bla(OXA-48)-like genes was constructed. The genetic context of the bla(OXA-48)-like gene in strain WCJ25 was investigated by inverse PCR using self-ligated AseI- or RsaI-restricted WCJ25 DNA fragments as template, while that in strain S4 was determined by PCR mapping using that in WCJ25 as template.ResultsA new bla(OXA-48) variant, designated bla(OXA-48b), with four silent nucleotide differences from the bla(OXA-48) (designated bla(OXA-48a)) found in the Enterobacteriaceae was identified in strain S4. Strain WCJ25 had a new bla(OXA-48)-like variant, bla(OXA-199), with five nucleotide differences from bla(OXA-48)a and bla(OXA-48b). The OXA-199 protein has three amino acid substitutions (H37Y, V44A and D153G) compared with OXA-48. Both bla(OXA-48b) and bla(OXA-199) were found adjacent to genes encoding a peptidase (indicated as orf), a protein of unknown function (sprT), an endonuclease I (endA), and a ribosomal RNA methyl transferase (rsmE) upstream and to transcriptional regulator gene lysR and an acetyl-CoA carboxylase-encoding gene downstream. In addition, the insertion sequence ISShes2 was found inserted downstream of bla(OXA-199) but not of bla(OXA-48b). The 26 bp sequences upstream and 63 bp downstream of bla(OXA-48a), bla(OXA-48b) and bla(OXA-199) were identical.Conclusionsbla(OXA-48a), bla(OXA-48b) and bla(OXA-199) might have a common origin, suggesting that the bla(OXA-48a) gene found in the Enterobacteriaceae could have originated from the chromosome of S. xiamenensis.

Project description:The emergence and spread of carbapenem-resistant Klebsiella pneumoniae (CRKP) is a serious medical problem worldwide. Acquired OXA-48-like carbapenemases encoded by plasmids are important causes of carbapenem resistance in K. pneumoniae. To explore the links between plasmids and bla OXA-48-like genes in K. pneumoniae, we systematically analyzed the variants of bla OXA-48-like plasmid replicon types, phylogenetic patterns, geographic distribution, conjugative transfer regions, and the genetic environments surrounding bla OXA-48-like of 191 bla OXA-48-like-harboring plasmids, which were identified from 4451 plasmids of K. pneumoniae downloaded from GenBank. Our results showed that seven different variants of bla OXA-48-like genes were identified from the 191 bla OXA-48-like-harboring plasmids in K. pneumoniae, with bla OXA-48, bla OXA-232, and bla OXA-181 being highly prevalent. In K. pneumoniae, bla OXA-48 was mainly carried by the composite transposon Tn1999.2 located on IncL/M-type conjugative plasmids, which were mainly geographically distributed in Switzerland, Germany, and China. In K. pneumoniae, the blaOXA-232 gene was mainly carried by 6.1-kb ColKP3-type mobilizable plasmids, which were mainly isolated in India. In K. pneumoniae, bla OXA-181 was mainly carried by a group of 50-kb ColKP3-IncX3 hybrid conjugative plasmids and a group of small ColKP3-type mobilizable plasmids with lengths of 5.9-9.3 kb, the former was sporadically discovered in China, South Korea, India, and Czech Republic, while the latter was almost all isolated in India. In addition, five bla OXA-245-harboring 65.9-kb IncL plasmids of K. pneumoniae isolated in Spain were found to have the genetic context of bla OXA-245 more complicated than that of bla OXA-48-harboring IncL/M-type plasmids, with two copies of IS1R inserted both upstream and downstream of bla OXA-245-lysR. These findings enhance our understanding of the genetic diversity of bla OXA-48-like-harboring plasmids in K. pneumoniae.

Project description:Aim of this study was to genetically characterize two carbapenemase-producing Escherichia coli strains obtained from a pediatric patient affected by diarrhea, expressing OXA-181 and/or NDM-5 type enzymes. The above microorganisms were collected in the same Desenzano hospital (Northern Italy) where the bla NDM- 5 gene was detected for the first time in Italy 3 years ago. One strain (5P), belonged to sequence type ST405/ST477 (according to Pasture/Oxford schemes) and serotype O102:H6. It was characterized by a 130562 bp multi-replicon plasmid IncFII/IncFIA/IncFIB (pVSI_NDM-5) enclosing two main antibiotic resistance islands: (i) ARI-I, 10030 bp in size, carried genes coding for β-lactam- (bla OXA- 1, bla CTX-M- 15), fluoroquinolone/aminoglycoside- (aac(6')-lb-cr) and phenicol- resistance (catB3), (ii) ARI-II, 15326 bp in size, carried genes coding for sulfonamide- (sul1), β-lactam- (bla NDM- 5, bla TEM- 1 B), phenicol- (catB3), trimethoprim- (dfrA17), antiseptic- (qacEΔ1), and aminoglycoside- (aadA5, rmtB) resistance. The other isolate (5M), belonged to sequence type ST2659/ST759 and serotype O50/02:H18, and carried four plasmids: a 153866 bp multi-replicon IncFII/IncFIA/IncFIB (pISV_IncFII_NDM-5), an 89866 bp IncI1 plasmid, a 51480 bp IncX3 plasmid (pISV_IncX3_OXA181), and a 41143 bp IncI plasmid (pISV_IncI_CMY-42). pISV_IncFII_NDM-5 carried two main antibiotic resistance islands: (i) ARI-III, 12220 bp in size, carried genes coding for β-lactam- (bla OXA- 1), fluoroquinolone/aminoglycoside- (aac(6')-lb-cr), tetracycline- (tet(B)) and phenicol- resistance (catB3, catA1), and ii) ARI-IV, 26527 bp in size, carried determinants coding for macrolide- (erm(B), mph(A)), sulfonamide- (sul1), beta-lactam- (bla NDM- 5, bla TEM- 1 B), trimethoprim- (dfrA14, dfrA12), antiseptic- (qacEΔ1), and aminoglycoside- resistance (aadA5). pISV_IncI_CMY-42 harbored the bla CMY- 42 gene coding for beta-lactam resistance, pISV_IncX3_OXA181 harbored genes encoding fluoroquinolone- (qnrS1) and beta-lactams- resistance (bla OXA- 181). In conclusion, the detection of two different NDM-5 E. coli strains from a pediatric patient with a history of travel to the Far East countries strongly highlight an increasing trend and risk of importation from such areas.

Project description:BackgroundThe spread of carbapenemase-producing Enterobacteriaceae (CPE) is a great problem of healthcare worldwide. Study of the spread for bla OXA-48-like genes coding epidemically significant carbapenemases among hospital pathogens is important for the regional and global epidemiology of antimicrobial resistance.MethodsAntibacterial resistant isolates of Klebsiella pneumoniae (n = 95) from 54 patients, P. mirabilis (n = 32) from 20 patients, Enterobacter aerogenes (n = 6) from four patients, and Enterobacter cloacae (n = 4) from four patients were collected from January, 2013 to October, 2014 in neurosurgical intensive care unit (ICU) of the Burdenko Neurosurgery Institute, Moscow. Characteristics of the isolates were done using susceptibility tests, PCR detection of the resistance genes, genotyping, conjugation, DNA sequencing, and bioinformatic analysis.ResultsMajor strains under study were multi drug resistant (MDR), resistant to three or more functional classes of drugs simultaneously-98.9 % K. pneumoniae, 100 % P. mirabilis, one E. aerogenes isolate, and one E. cloacae isolate. Molecular-genetic mechanism of MDR in K. pneumoniae and P. mirabilis isolates were based on carrying of epidemic extended-spectrum beta-lactamase bla CTX-M-15 gene (87.2 and 90.6 % accordingly), carbapenemase bla OXA-48-like gene (55.3 and 23.3 % accordingly), and class 1 (54.8 and 31.3 % accordingly) and class 2 (90.6 % P. mirabilis) integrons. The bla OXA-48-like-positive K. pneumoniae were collected during whole two-year surveillance period, while P. mirabilis and Enterobacter spp. carrying bla OXA-48-like genes were detected only after four and 18 months after the research start, respectively. The bla OXA-48-like gene acquisition was shown for P. mirabilis isolates collected from five patients and for E. cloacae isolate collected from one patient during their stay in the ICU, presumably from bla OXA-48-like-positive K. pneumoniae. The source of the bla OXA-244 gene acquired by E. aerogenes isolates and the time of this event were not recognized.ConclusionsThe expanding of CPE in the surveyed ICU was associated with the spread of bla OXA-48 and bla OXA-244 carbapenemase genes documented not only among K. pneumoniae, well-known bacterial host for such genes, but among P. mirabilis, E. aerogenes, and E. cloacae.

Project description:Purpose:The emergence of multidrug-resistant Klebsiella pneumoniae (K. pneumoniae) is associated with the acquisition of multiple carbapenemases. Their clonal spread is a worldwide concern due to their critical role in nosocomial infections. Therefore, the identification of high-risk clones with antibiotic resistance genes is very crucial for controlling its global spread. Materials and Methods:A total of 227 K. pneumoniae strains collected during April 2018 to November 2019 were confirmed by PCR. Carbapenemases and extended-spectrum ?-lactamases (ESBL) were detected phenotypically. Confirmation of carbapenemases was carried out by PCR and Sanger sequencing. The clonal lineages were assigned to selected isolates by multilocus sequence typing (MLST), and the plasmid analysis was done by PCR-based detection of the plasmid replicon typing. Results:Of the total K. pneumoniae, 117 (51.5%) were carbapenem resistant (CRKP) and 140 (61.7%) were identified as ESBL producers. Intermediate to high resistance was detected in the tested ?-lactam drugs while polymyxin-B and tigecycline were found to be susceptible. Among CRKP, 91 (77.8%) isolates were detected as carbapenemase producing, while 55 (47%) were positive for bla NDM-1 23.9% (n=28), bla OXA-48 22.2% (n=26) and bla VIM 0.85% (n=1) while 12.7% (n=7) carried both bla NDM-1 and bla OXA-48 genes. The CRKP coharboring bla NDM-1 and bla OXA-48 genes (n=7) were positive for bla CTX-M bla SHV (n=3), bla SHV (n=1) and bla CTX-M (n=3). The novel CRKP with the coexistence of bla NDM-1, bla OXA-48, bla CTX-M and bla SHV genes were associated with the high-risk clone ST147 (n=5) and ST11 (n=2). The assigned replicon types were IncL/M, IncFII, IncA/C and IncH1. Conclusion:This is the first report of the coexistence of bla NDM-1, bla OXA-48, bla CTX-M and bla SHV genes on a high-risk lineage ST147 from Pakistan. This study highlights the successful dissemination of carbapenemase resistance genes in the high-risk clones that emphasizes the importance of monitoring and controlling the spread of these diverse clones globally.

Dataset Information

IS1R-mediated plasticity of IncL/M plasmids leads to the insertion of bla OXA-48 into the Escherichia coli Chromosome.

Publications

IS1R-mediated plasticity of IncL/M plasmids leads to the insertion of bla OXA-48 into the Escherichia coli Chromosome.

Similar Datasets

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