Molecular Characterization of an IncFIIk Plasmid Co-harboring blaIMP–26 and tet(A) Variant in a Clinical Klebsiella pneumoniae Isolate

Yao, Hong; Cheng, Jing; Li, Aijuan; Yu, Ran; Zhao, Wenbo; Qin, Shangshang; Du, Xiang-Dang

doi:10.3389/fmicb.2020.01610

Cited by 15 publications

(15 citation statements)

References 31 publications

(39 reference statements)

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“…The FII plasmid pIMP-KP-13-9 was 112,209 bp long with an average GC content of 51.19% and encoding 138 predicted ORFs ( Figure 1B ). This plasmid showed 98.94% nucleotide identity and 84% query coverage with pIMP1572 (MH464586), a plasmid carrying both bla IMP – 26 and tet(A) variants ( Yao et al, 2020 ). Different from the plasmid pIMP1572, a Tn 1721 -like transposon structure carrying the tet (A) variant which is responsible for tigecycline was absent in pIMP-KP-13-9.…”

Section: Resultsmentioning

confidence: 99%

Molecular Characterization of blaIMP–4-Carrying Enterobacterales in Henan Province of China

et al. 2021

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Carbapenem-resistant Enterobacterales (CRE) pose a serious threat to clinical management and public health. We investigated the molecular characteristics of 12 IMP-4 metallo-β-lactamase-producing strains, namely, 5 Enterobacter cloacae, 3 Escherichia coli, 2 Klebsiella pneumoniae, and 2 Citrobacter freundii. These strains were collected from a tertiary teaching hospital in Zhengzhou from 2013 to 2015. The minimum inhibitory concentration (MIC) results showed that each blaIMP–4-positive isolate was multidrug-resistant (MDR) but susceptible to colistin. All of the E. coli belonged to ST167, two C. freundii isolates belonged to ST396, and diverse ST types were identified in E. cloacae and K. pneumoniae. S1-PFGE, Southern blotting, and PCR-based replicon typing assays showed that the blaIMP–4-carrying plasmids ranged from ∼52 to ∼360 kb and belonged to FII, FIB, HI2/HI2A, and N types. N plasmids were the predominant type (8/12, 66.7%). Plasmid stability testing indicated that the blaIMP–4-carrying N-type plasmid is more stable than the other types of plasmids. Conjugative assays revealed that three of the blaIMP–4-carrying N plasmids were transferrable. Complete sequence analysis of a representative N type (pIMP-ECL14–57) revealed that it was nearly identical to pIMP-FJ1503 (KU051710) (99% nucleotide identity and query coverage), an N-type blaIMP–4-carrying epidemic plasmid in a C. freundii strain. PCR mapping indicated that a transposon-like structure [IS6100-mobC-intron (K1.pn.I3)-blaIMP–4-IntI1-IS26] was highly conserved in all of the N plasmids. IS26 involved recombination events that resulted in variable structures of this transposon-like module in FII and FIB plasmids. The blaIMP–4 gene was captured by a sul1-type integron In1589 on HI2/HI2A plasmid pIMP-ECL-13–46.

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Section: Resultsmentioning

confidence: 99%

Molecular Characterization of blaIMP–4-Carrying Enterobacterales in Henan Province of China

et al. 2021

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“…In addition, tet (A) mutation occurring under selective pressure may lead to tigecycline treatment failure. Zhang et al (2019) reported the coexistence of mcr-1 and the tet (A) variant on the same plasmid from a K. pneumoniae isolate in human gut, and Yao et al (2020) also reported an IncFII plasmid co-harboring bla IMP– 26 and tet (A) variant in a clinical K. pneumoniae isolate. It seems that tet (A) mutants can not only occur in bla KPC– 2 -carrying plasmids, but also form fusion plasmids with other carbapenem resistance genes and mcr gene, which will cause a higher transmission risk of simultaneous resistance to carbapenem, colistin and tigecycline.…”

Section: Discussionmentioning

confidence: 97%

“…Multiple types of tet (A)-bearing plasmids were retrieved from the NCBI GenBank database, and circular comparison analysis revealed that they have some similar structures, suggesting that genetic exchange and recombination among different types of tet (A)-bearing plasmids have occurred ( Ribera et al, 2003 ; Szmolka et al, 2015 ; Yao et al, 2020 ). In the three tet (A)-bearing plasmids obtained in this study, one class 1 integron containing multiple ARGs, including tet (A), was detected.…”

Section: Discussionmentioning

confidence: 99%

The Plasmid-Borne tet(A) Gene Is an Important Factor Causing Tigecycline Resistance in ST11 Carbapenem-Resistant Klebsiella pneumoniae Under Selective Pressure

Zhu

Chen

et al. 2021

Front. Microbiol.

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The emergence and prevalence of tigecycline-resistant Klebsiella pneumoniae have seriously compromised the effectiveness of antimicrobial agents in the treatment of infections. To explore the role of the plasmid-borne tet(A) gene in tigecycline resistance in carbapenem-resistant K. pneumoniae (CRKP), a total of 63 CRKP isolates were collected from a tertiary hospital in Hangzhou, China. The minimum inhibitory concentration (MIC) of tigecycline, mutation rate of tet(A) gene, genetic surroundings of tet(A)-carrying transmissible plasmid and the contribution of tet(A) mutation to tigecycline resistance were analyzed using antimicrobial susceptibility test, whole-genome sequencing, tigecycline resistance evolution experiment, and plasmid conjugation experiment. Our results showed that 52.4% (33 isolates) of the test isolates carried the tet(A) gene; among them, 75.8% (25 isolates) exhibited a tigecycline non-susceptible phenotype (MIC = 4 mg/L). Three clonal groups (cluster I, cluster II, and cluster III) were identified in these tet(A)-bearing isolates. All 17 isolates belonged to serotype KL21 (cluster I), which differed by only 13 SNPs, suggesting a clonal spread of tet(A)-positive ST11 K. pneumoniae with serotype KL21 occurred in the sampling hospital. The induction of tigecycline resistance experiments showed that 71.4% of strains evolved tet(A) mutations and developed a high-level tigecycline resistance. Eight amino acid substitutions were identified in these mutants. The most common amino acid substitution was A370V, followed by S251A and G300E. Twelve isolates carrying tet(A) mutants succeeded in the filter mating experiment with a conjugation efficiency of 10–3–10–8. Tigecycline MICs in E. coli EC600 transconjugants with a mutated tet(A) were 2 to 8-fold higher than those in E. coli EC600 transconjugants with a wild-type tet(A). One ColRNAI/IncFII type and two IncFII type tet(A)-bearing conjugative plasmids were identified in this study, including a class 1 integron containing multiple antibiotic resistance genes, i.e., tet(A), qnrS1, blaLAP–2, catA2, sul2, and dfrA14. Our study revealed the wide-spread situation of plasmid-borne tet(A) gene in clinical CRKP, and mutation of tet(A) is a potential driven force that lead to tigecycline resistance.

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“…Hypothetical pKpQIL_019 protein gene was identified on plasmid pKpQIL next to the Tn4401 transposon between the transposase and resolvase genes [ 60 , 61 ]. The Tn4401 transposon has also been identified in other plasmids [ 56 ], specifically in the plasmid IncFIIK [ 62 , 63 ], IncN [ 64 ] and ColE [ 65 , 66 ], IncI2 [ 67 , 68 ], and IncX3 [ 69 , 70 ]. A thorough analysis of the pKpQIL sequences in the BLAST databases revealed that the p019 sequence responsible to produce the cleavage product of the hypothetical pKpQIL_019 protein was always identified in the bla KPC gene.…”

Section: Resultsmentioning

confidence: 99%

Carbapenemase Producing Klebsiella pneumoniae (KPC): What Is the Best MALDI-TOF MS Detection Method

et al. 2021

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Klebsiella pneumoniae carbapenemase (KPC)-producing bacteria is a group of highly dangerous antibiotic resistant Gram-negative Enterobacteriaceae. They cause infections associated with significant morbidity and mortality. Therefore, the rapid detection of KPC-producing bacteria plays a key role in clinical microbiology. Matrix assisted laser desorption/ionization time-of- flight (MALDI-TOF) is a rapidly evolving technology that finds application in various clinical, scientific, and industrial disciplines. In the present study, we demonstrated three different procedures of carbapenemase-producing K. pneumoniae (KPC) detection. The most basic model of MALDI-TOF instrument MS Microflex LT was used, operating in the linear ion-positive mode, commonly used in modern clinical laboratories. The first procedure was based on indirect monitoring of carbapenemase production with direct detection of hydrolyzed carbapenem antibiotic degradation products in the mass spectrum. The second procedure was based on direct detection of blaKPC accompanying peak with an 11,109 Da in the mass spectrum of carbapenemase-producing K. pneumoniae (KPC), which represents the cleaved protein (pKpQIL_p019) expressed by pKpQIL plasmid. In addition, several unique peaks were detected in the carbapenemase-producing K. pneumoniae (KPC) mass spectrum. The third procedure was the identification of carbapenemase-producing K. pneumoniae (KPC) based on the protein fingerprint using local database created from the whole mass spectra. By comparing detection procedures, we determined that the third procedure was very fast and relatively easy. However, it requires previous verification of carbapenemase-producing K. pneumoniae (KPC) using other methods as genetic blaKPC identification, detection of carbapenem degradation products, and accompanying peak with 11,109 Da, which represents cleaved pKpQIL_p019 protein expressed by pKpQIL plasmid. Detection of carbapenemase-producing K. pneumoniae using MALDI-TOF provides fast and accurate results that may help to reduce morbidity and mortality in hospital setting when applied in diagnostic situations.

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Molecular Characterization of an IncFIIk Plasmid Co-harboring blaIMP–26 and tet(A) Variant in a Clinical Klebsiella pneumoniae Isolate

Cited by 15 publications

References 31 publications

Molecular Characterization of blaIMP–4-Carrying Enterobacterales in Henan Province of China

Molecular Characterization of blaIMP–4-Carrying Enterobacterales in Henan Province of China

The Plasmid-Borne tet(A) Gene Is an Important Factor Causing Tigecycline Resistance in ST11 Carbapenem-Resistant Klebsiella pneumoniae Under Selective Pressure

Carbapenemase Producing Klebsiella pneumoniae (KPC): What Is the Best MALDI-TOF MS Detection Method

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