Dataset Information

Novel chromosomal insertions of ISEcp1-bla_CTX-M-15 and diverse antimicrobial resistance genes in Zambian clinical isolates of Enterobacter cloacae and Escherichia coli.

ABSTRACT:

Background

The epidemiology of extended-spectrum β-lactamases (ESBLs) has undergone dramatic changes, with CTX-M-type enzymes prevailing over other types. bla_CTX-M genes, encoding CTX-M-type ESBLs, are usually found on plasmids, but chromosomal location is becoming common. Given that bla_CTX-M-harboring strains often exhibit multidrug resistance (MDR), it is important to investigate the association between chromosomally integrated bla_CTX-M and the presence of additional antimicrobial resistance (AMR) genes, and to identify other relevant genetic elements.

Methods

A total of 46 clinical isolates of cefotaxime-resistant Enterobacteriaceae (1 Enterobacter cloacae, 9 Klebsiella pneumoniae, and 36 Escherichia coli) from Zambia were subjected to whole-genome sequencing (WGS) using MiSeq and MinION. By reconstructing nearly complete genomes, bla_CTX-M genes were categorized as either chromosomal or plasmid-borne.

Results

WGS-based genotyping identified 58 AMR genes, including four bla_CTX-M alleles (i.e., bla_CTX-M-14, bla_CTX-M-15, bla_CTX-M-27, and bla_CTX-M-55). Hierarchical clustering using selected phenotypic and genotypic characteristics suggested clonal dissemination of bla_CTX-M genes. Out of 45 bla_CTX-M gene-carrying strains, 7 harbored the gene in their chromosome. In one E. cloacae and three E. coli strains, chromosomal bla_CTX-M-15 was located on insertions longer than 10 kb. These insertions were bounded by ISEcp1 at one end, exhibited a high degree of nucleotide sequence homology with previously reported plasmids, and carried multiple AMR genes that corresponded with phenotypic AMR profiles.

Conclusion

Our study revealed the co-occurrence of ISEcp1-bla_CTX-M-15 and multiple AMR genes on chromosomal insertions in E. cloacae and E. coli, suggesting that ISEcp1 may be responsible for the transposition of diverse AMR genes from plasmids to chromosomes. Stable retention of such insertions in chromosomes may facilitate the successful propagation of MDR clones among these Enterobacteriaceae species.

SUBMITTER: Shawa M

PROVIDER: S-EPMC8111917 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The worldwide dissemination of CTX-M type β-lactamases is a threat to human health. Previously, we have reported the spread of bla(CTX-M-15) gene in different clinical strains of Enterobacteriaceae from the hospital settings of Aligarh in north India. In view of the varying resistance pattern against cephalosporins and other β-lactam antibiotics, we intended to understand the correlation between MICs and catalytic activity of CTX-M-15. In this study, steady-state kinetic parameters and MICs were determined on E. coli DH5α transformed with bla(CTX-M-15) gene that was cloned from Enterobacter cloacae (EC-15) strain of clinical background. The effect of conventional β-lactamase inhibitors (clavulanic acid, sulbactam and tazobactam) on CTX-M-15 was also studied. We have found that tazobactam is the best among these inhibitors against CTX-M-15. The inhibition characteristic of tazobactam is defined by its very low IC(50) value (6 nM), high affinity (K(i) = 0.017 µM) and better acylation efficiency (k(+2)/K' = 0.44 µM(-1)s(-1)). It forms an acyl-enzyme covalent complex, which is quite stable (k(+3) = 0.0057 s(-1)). Since increasing resistance has been reported against conventional β-lactam antibiotic-inhibitor combinations, we aspire to design a non-β-lactam core containing β-lactamase inhibitor. For this, we screened ZINC database and performed molecular docking to identify a potential non-β-lactam based inhibitor (ZINC03787097). The MICs of cephalosporin antibiotics in combination with this inhibitor gave promising results. Steady-state kinetics and molecular docking studies showed that ZINC03787097 is a reversible inhibitor which binds non-covalently to the active site of the enzyme through hydrogen bonds and hydrophobic interactions. Though, it's IC(50) (180 nM) is much higher than tazobactam, it has good affinity for CTX-M-15 (K(i) = 0.388 µM). This study concludes that ZINC03787097 compound can be used as seed molecule to design more efficient non-β-lactam containing β-lactamase inhibitor that could evade pre-existing bacterial resistance mechanisms.

Project description:IncX3 and IncL plasmids have been named as catalysts advancing dissemination of blaOXA-181 and blaOXA-48 genes. However, their impact on the performance of host cells is vastly understudied. Genetic characteristics of blaOXA-48- and blaOXA-181-containing Klebsiella pneumoniae (EFN299), Klebsiella quasipneumoniae (EFN262), and Enterobacter cloacae (EFN743) isolated from clinical samples in a Ghanaian hospital were investigated by whole-genome sequencing. Transfer of plasmids by conjugation and electroporation, plasmid stability, fitness cost, and genetic context of blaOXA-48, blaOXA-181, and blaDHA-1 were assessed. blaOXA-181 was carried on two IncX3 plasmids, an intact 51.5-kb IncX3 plasmid (p262-OXA-181) and a 45.3-kb IncX3 plasmid (p743-OXA-181) without replication protein sequence. The fluoroquinolone-resistant gene qnrS1 region was also excised, and unlike in p262-OXA-181, the blaOXA-181 drug-resistant region was not found on a composite transposon. blaOXA-48 was carried on a 74.6-kb conjugative IncL plasmid with unknown ~10.9-kb sequence insertion. This IncL plasmid proved to be highly transferable, with a conjugation efficiency of 1.8 × 10-2. blaDHA-1 was present on an untypeable 22.2 kb genetic structure. Plasmid stability test revealed plasmid loss rate between 4.3% and 12.4%. The results also demonstrated that carriage of IncX3-blaOXA-181 or IncL-blaOXA-48 plasmids was not associated with any fitness defect, but rather an enhanced competitive ability of host cells. This study underscores the significant contribution of IncX3 and IncL plasmids in the dissemination of resistance genes and their efficient transfer calls for regular monitoring to control the expansion of resistant strains. IMPORTANCE The growing rate of antibiotic resistance is an important global health threat. This threat is exacerbated by the lack of safe and potent alternatives to carbapenems in addition to the slow developmental process of newer and effective antibiotics. Infections by carbapenem-resistant Gram-negative bacteria are becoming almost untreatable, leading to poor clinical outcomes and high mortality rates. OXA-48-like carbapenemases are one of the most widespread carbapenemases accounting for resistance among Enterobacteriaecae. We characterized OXA-48- and OXA-181-producing Enterobacteriaecae to gain insights into the genetic basis and mechanism of resistance to carbapenems. Findings from the study showed that the genes encoding these enzymes were carried on highly transmissible plasmids, one of which had sequences absent in other similar plasmids. This implies that mobile genetic elements are important players in the dissemination of resistance genes. Further characterization of this plasmid is warranted to determine the role of this sequence in the spread of resistance genes.

Dataset Information

Novel chromosomal insertions of ISEcp1-bla_CTX-M-15 and diverse antimicrobial resistance genes in Zambian clinical isolates of Enterobacter cloacae and Escherichia coli.

Background

Methods

Results

Conclusion

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OmicsDI is part of the ELIXIR infrastructure

Tweets

Dataset Information

Novel chromosomal insertions of ISEcp1-blaCTX-M-15 and diverse antimicrobial resistance genes in Zambian clinical isolates of Enterobacter cloacae and Escherichia coli.

Background

Methods

Results

Conclusion

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Novel chromosomal insertions of ISEcp1-bla_CTX-M-15 and diverse antimicrobial resistance genes in Zambian clinical isolates of Enterobacter cloacae and Escherichia coli.