BACKGROUND: VIM metallo-β-lactamases are enzymes characterized by the ability to hydrolyze all β-lactams. Usually, blaVIM-like genes are carried by class 1 integrons. In the Czech Republic, only sporadic cases of VIM-producing Enterobacterales have been reported in which those isolates carried the VIM-1 carbapenemase-encoding integron In110. However, during 2019-2020, an increased number was reported. Therefore, the aim of the current study was to characterize the genetic elements involved in the increased spread of blaVIM genes. MATERIALS AND METHODS: 32 VIM-producing Enterobacterales collected between 2019 and 2020 were subjected to: antimicrobial susceptibility testing, integron analysis, and short reads sequencing. Based on the results, 19 isolates were selected as representative and sequenced using Sequel I platform. RESULTS: The 32 VIM-producing isolates exhibited variations in the MICs of carbapenems. Based on short-read data, 26 of the 32 sequenced isolates harbored the blaVIM-1 allele while six isolates carried the blaVIM-4 gene. The most prevalent was the In110 integron (n = 24) and two isolates carried the In4873 class 1 integron. The blaVIM-4 allele was identified in class 1 integrons In1174 (n = 3), In416 (n = 1), In2143 (n = 1) and In2150. Long reads sequencing revealed that the blaVIM was carried by: pKPC-CAV1193-like (n = 6), HI1 (pNDM-CIT; n = 4), HI2 (n = 3), FIB (pECLA; n = 2) and N (n = 1) incompatibility groups. Two blaVIM-carrying plasmids could not be typed by the database, while another one was integrated into the chromosome. CONCLUSION: We observed the spread of VIM-encoding integrons, mainly of In110, among Enterobacterales isolated from Czech hospitals, but also an increased number of novel elements underlining the ongoing evolution.

• Publikační typ
• časopisecké články MeSH

The aim of this study was to report the characterization of the first mcr-positive Enterobacterales isolated from Czech hospitals. In 2019, one Citrobacter freundii and four Enterobacter isolates were recovered from Czech hospitals. The production of carbapenemases was examined by a matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) imipenem hydrolysis assay. Additionally, bacteria were screened for the presence of carbapenemase-encoding genes and plasmid-mediated colistin resistance genes by PCR. To define the genetic units carrying mcr genes, the genomic DNAs of mcr-carrying clinical isolates were sequenced on the PacBio Sequel I platform. Results showed that all isolates carried blaVIM- and mcr-like genes. Analysis of whole-genome sequencing (WGS) data revealed that all isolates carried mcr-9-like alleles. Furthermore, the three sequence type 106 (ST106) Enterobacter hormaechei isolates harbored the blaVIM-1 gene, while the ST764 E. hormaechei and ST95 C. freundii included blaVIM-4 Analysis of plasmid sequences showed that, in all isolates, mcr-9 was carried on IncHI2 plasmids. Additionally, at least one multidrug resistance (MDR) region was identified in each mcr-9-carrying IncHI2 plasmid. The blaVIM-4 gene was found in the MDR regions of p48880_MCR_VIM and p51929_MCR_VIM. In the three remaining isolates, blaVIM-1 was localized on plasmids (∼55 kb) exhibiting repA-like sequences 99% identical to the respective gene of pKPC-CAV1193. In conclusion, to the best of our knowledge, these 5 isolates were the first mcr-9-positive bacteria of clinical origin identified in the Czech Republic. Additionally, the carriage of the blaVIM-1 on pKPC-CAV1193-like plasmids is described for the first time. Thus, our findings underline the ongoing evolution of mobile elements implicated in the dissemination of clinically important resistance determinants.IMPORTANCE Infections caused by carbapenemase-producing bacteria have led to the revival of polymyxins as the "last-resort" antibiotic. Since 2016, several reports describing the presence of plasmid-mediated colistin resistance genes, mcr, in different host species and geographic areas were published. Here, we report the first detection of Enterobacterales carrying mcr-9-like alleles isolated from Czech hospitals in 2019. Furthermore, the three ST106 Enterobacter hormaechei isolates harbored blaVIM-1, while the ST764 E. hormaechei and ST95 Citrobacter freundii isolates included blaVIM-4 Analysis of WGS data showed that, in all isolates, mcr-9 was carried on IncHI2 plasmids. blaVIM-4 was found in the MDR regions of IncHI2 plasmids, while blaVIM-1 was localized on pKPC-CAV1193-like plasmids, described here for the first time. These findings underline the ongoing evolution of mobile elements implicated in dissemination of clinically important resistance determinants. Thus, WGS characterization of MDR bacteria is crucial to unravel the mechanisms involved in dissemination of resistance mechanisms.

• MeSH
• antibakteriální látky farmakologie   terapeutické užití   MeSH
• bakteriální proteiny genetika   MeSH
• beta-laktamasy genetika   MeSH
• Enterobacter genetika   izolace a purifikace   MeSH
• lidé MeSH
• nemocnice MeSH
• plazmidy genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH

Carbapenemase-producing Enterobacteriaceae and Pseudomonas spp. are increasingly reported in many countries all over the world. Due to the resistance of those bacteria to almost all antibiotics (e.g.beta-lactams, aminoglycosides, fluoroquinolones),treatment options are seriously limited. In the Czech Republic, the incidence of carbapenemase-producing Enterobacteriaceae seems to be low, restricted to only three cases detected between 2009 and 2010.Here, we describe molecular typing of 15 carbapenemase-producing Klebsiella pneumoniae isolates identified in the Czech Republic during 2011. Five VIM-1-producing isolates belonging to sequence type (ST)11 and one VIM-4-producing isolate of ST1029 have been detected. blaVIM-1 and blaVIM-4 as a part of class 1 integrons were chromosomally located or carried by a plasmid belonging to A/C replicon type (blaVIM-4). KPC-3-producing isolates of ST512, recovered from six patients, caused an outbreak. Three more isolates producing KPC-2 enzyme belonged to ST258. Both blaKPCgenes were part of the Tn4401a transposon carried on plasmids of the pKpQIL type. The isolates were resistant to all antibiotics tested except colistin and/or gentamicin.Four of these 15 strains were recovered from patients repatriated to the Czech Republic from Greece and Italy. This is the first report of outbreaks caused by carbapenemase-producing Enterobacteriaceae in the Czech Republic.

• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální proteiny metabolismus   MeSH
• beta-laktamasy metabolismus   MeSH
• beta-laktamová rezistence MeSH
• cestování MeSH
• incidence MeSH
• infekce bakteriemi rodu Klebsiella diagnóza   enzymologie   epidemiologie   MeSH
• integrony genetika   MeSH
• karbapenemy farmakologie   MeSH
• Klebsiella pneumoniae klasifikace   účinky léků   genetika   izolace a purifikace   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• mnohočetná bakteriální léková rezistence MeSH
• molekulární typizace MeSH
• plazmidy genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Itálie MeSH
• Řecko MeSH

BACKGROUND: the co-production of carbapenemases and mcr-genes represents a worrisome event in the treatment of Enterobacteriaceae infections. The aim of the study was to characterize the genomic features of two clinical Enterobacter cloacae complex (ECC) isolates, co-producing VIM and MCR enzymes, in Italy. METHODS: species identification and antibiotic susceptibility profiling were performed using MALDI-TOF and broth microdilution methods, respectively. Transferability of the blaVIM- and mcr- type genes was verified through conjugation experiment. Extracted DNA was sequenced using long reads sequencing technology on the Sequel I platform (PacBio). RESULTS: the first isolate showed clinical resistance against ertapenem yet was colistin susceptible (EUCAST 2020 breakpoints). The mcr-9.2 gene was harbored on a conjugative IncHI2 plasmid, while the blaVIM-1 determinant was harbored on a conjugative IncN plasmid. The second isolate, resistant to both carbapenems and colistin, harbored: mcr-9 gene and its two component regulatory genes for increased expression on the chromosome, mcr-4.3 on non-conjugative (yet co-transferable) ColE plasmid, and blaVIM-1 on a non-conjugative IncA plasmid. CONCLUSIONS: to our knowledge, this is the first report of co-production of VIM and MCR in ECC isolates in Italy.

• Publikační typ
• časopisecké články MeSH

A VIM-1-producing ST92 Enterobacter cloacae was isolated in a Czech hospital. blaVIM-1 was part of the class 1 integron In110 carried by a Tn1721-like transposon. Tn1721-like was located on a ColE1-like plasmid, pEncl-30969cz (33,003 bp). Target site duplications at the boundaries of Tn1721-like suggested its transposition into the pEncl-30969cz backbone.

• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální proteiny genetika   MeSH
• beta-laktamasy genetika   MeSH
• beta-laktamová rezistence genetika   MeSH
• DNA bakterií analýza   genetika   MeSH
• Enterobacter cloacae genetika   MeSH
• enterobakteriální infekce mikrobiologie   MeSH
• integrony genetika   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• plazmidy genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Sequence type 11 Klebsiella pneumoniae, coproducing NDM-1 and VIM-1 metallo-β-lactamases, were isolated in a Greek hospital. blaNDM-1 was part of a Tn125 derivative, located on an ~90-kb plasmid similar to the NDM-1-encoding plasmid pB-3002cz. blaVIM-1 was located in an In-e541-like integron, carried on a multireplicon (IncA/C and IncR) plasmid of ~180kb.

• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• beta-laktamasy genetika   metabolismus   MeSH
• infekce bakteriemi rodu Klebsiella farmakoterapie   mikrobiologie   MeSH
• karbapenemy farmakologie   MeSH
• Klebsiella pneumoniae klasifikace   genetika   izolace a purifikace   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• nemocnice MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Řecko MeSH

The complete nucleotide sequences of three multidrug resistance (MDR) IncA/C-like plasmids from Enterobacteriaceae isolates carrying the VIM-type carbapenemase-encoding integrons In4863 (blaVIM-19-aacA7-dfrA1-ΔaadA1-smr2) or In4873 (blaVIM-1-aacA7-dfrA1-ΔaadA1-smr2) were determined, which are the first In416-like elements identified in Greece. Plasmids pKP-Gr642 and pKP-Gr8143 were from Klebsiella pneumoniae ST383 isolates, whereas plasmid pEcl-Gr4873 was from an Enterobacter cloacae ST88 isolate. Sequencing showed that pKP-Gr642 (162787bp) and pKP-Gr8143 (154395bp) consisted of the type 1 IncA/C2 conserved backbone, the blaCMY-2-like gene-containing region, and the ARI-B (with the sul2 gene) and ARI-A (with a class 1 integron) resistance islands, like the plasmid pUMNK88_161 from the USA. The third plasmid, pEcl-Gr4873 (153958bp), exhibited extensive similarity with the type 2 IncA/C2 plasmid pR55 from France. pEcl-Gr4873 carried only one resistance island of a hybrid transposon structure inserted in a different location to ARI-A in type 1 A/C2 plasmids. In all three plasmids, the In416-like integrons In4863 or In4873 were identified within non-identical class II transposon structures. All three In416-like-carrying regions presented significant similarities with the MDR region of the IncA/C2 plasmid pCC416 from Italy, carrying the prototype In416 integron (blaVIM-4-aacA7-dfrA1-ΔaadA1-smr2). These findings provided the basis for speculations regarding the evolution of IncA/C2 plasmids with In416-like integrons, and confirmed the rapid evolution of some IncA/C2 plasmid lineages. Considering the broad host range of IncA/C2 molecules, it seems that pKP-Gr642, pKP-Gr8143 and pEcl-Gr4873 plasmids might support the diffusion of In416-like integrons among Enterobacteriaceae.

• MeSH
• beta-laktamasy genetika   MeSH
• Enterobacter cloacae enzymologie   genetika   izolace a purifikace   MeSH
• enterobakteriální infekce mikrobiologie   MeSH
• integrony * MeSH
• Klebsiella pneumoniae enzymologie   genetika   izolace a purifikace   MeSH
• lidé MeSH
• molekulární sekvence - údaje MeSH
• plazmidy * MeSH
• pořadí genů MeSH
• sekvenční analýza DNA MeSH
• sekvenční homologie MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Řecko MeSH

Background: VIM (Verona Integron-encoded Metallo-beta-lactamase) is a member of the Metallo-Beta-Lactamases (MBLs), and is able to hydrolyze all beta-lactams antibiotics, except for monobactams, and including carbapenems. Here we characterize a VIM-producing IncA plasmid isolated from a clinical ST69 Escherichia coli strain from an Italian Long-Term Care Facility (LTCF) inpatient. Methods: An antimicrobial susceptibility test and conjugation assay were carried out, and the transferability of the blaVIM-type gene was confirmed in the transconjugant. Whole-genome sequencing (WGS) of the strain 550 was performed using the Sequel I platform. Genome assembly was performed using "Microbial Assembly". Genomic analysis was conducted by uploading the contigs to ResFinder and PlasmidFinder databases. Results: Assembly resulted in three complete circular contigs: the chromosome (4,962,700 bp), an IncA plasmid (p550_IncA_VIM_1; 162,608 bp), harboring genes coding for aminoglycoside resistance (aac(6')-Ib4, ant(3″)-Ia, aph(3″)-Ib, aph(3')-XV, aph(6)-Id), beta-lactam resistance (blaSHV-12, blaVIM-1), macrolides resistance (mph(A)), phenicol resistance (catB2), quinolones resistance (qnrS1), sulphonamide resistance (sul1, sul2), and trimethoprim resistance (dfrA14), and an IncK/Z plasmid (p550_IncB_O_K_Z; 100,306 bp), free of antibiotic resistance genes. Conclusions: The increase in reports of IncA plasmids bearing different antimicrobial resistance genes highlights the overall important role of IncA plasmids in disseminating carbapenemase genes, with a preference for the blaVIM-1 gene in Italy.

• Publikační typ
• časopisecké články MeSH

The objective of this study was to assess the susceptibility of cefiderocol against multidrug-resistant carbapenemase-producing and nonproducing bacteria. The panel comprised 182 isolates of the order Enterobacterales, and 40 strains of Pseudomonas aeruginosa. Antimicrobial susceptibility testing has been performed using broth microdilution method according to the European Committee on Antimicrobial Susceptibility Testing recommendations. Mass spectrometry matrix-assisted laser desorption/ionization-time of flight mass spectrometry and carbapenemase-producing test were used to verify the presence of carbapenemases in clinical isolates. The genetic expression of single carbapenemases (blaKPC, blaOXA-48, blaNDM, blaVIM, blaIMP, blaGES) was determined by real-time polymerase chain reaction. Cefiderocol exhibited a good activity against the majority of strains tested in this study. Altogether, growth of 81.9% (n = 149) strains of the order Enterobacterales and 77.5% (n = 31) of P. aeruginosa isolates were inhibited at minimal inhibitory concentration (MIC) ≤2 mg/L. Values MIC50/MIC90 were 0.5/8 mg/L for enterobacteria, and 1/8 mg/L for P. aeruginosa. One isolate (Klebsiella pneumoniae) harboring two carbapenemases (blaOXA-48, blaNDM) had cefiderocol MIC 0.5 mg/L. In enterobacteria resistant to cefiderocol, blaNDM carbapenemase prevailed (43.3%, n = 29), followed by blaOXA-48 (31.3%, n = 21) and blaKPC (4.5%, n = 3). blaIMP (n = 8) and blaVIM (n = 1) metallo-β-lactamases dominated in cefiderocol-resistant P. aeruginosa (n = 9) isolates. Very good susceptibility (100%) to this drug showed blaGES-positive strains of P. aeruginosa (n = 8) and isolates resistant to meropenem without confirmed carbapenemase gene (n = 10). In this study, cefiderocol demonstrated potent activity against important nosocomial pathogens, therefore, therapeutic options of this drug against multidrug-resistant bacteria should be considered.

• MeSH
• antibakteriální látky * farmakologie   MeSH
• beta-laktamasy genetika   metabolismus   MeSH
• cefiderokol MeSH
• Enterobacteriaceae genetika   metabolismus   MeSH
• karbapenemy * farmakologie   MeSH
• mikrobiální testy citlivosti MeSH
• Pseudomonas aeruginosa MeSH
• Publikační typ
• časopisecké články MeSH

Fosfomycin (FOS) is an effective antibiotic against multidrug-resistant Enterobacterales, but its effectiveness is reducing. Little is known on the current prevalence of FosA enzymes in low-risk pathogens, such as Citrobacter freundii. The aim of the study was the molecular characterization of a carbapenemase- and FosA-producing C. freundii collected in Italy. AK867, collected in 2023, showed an XDR profile, retaining susceptibility only to colistin. AK867 showed a FOS MIC >128 mg/L by ADM. Based on WGS, AK867 belonged to ST116 and owned a wide resistome, including fosA3, blaKPC-2, and blaVIM-1. fosA3 was carried by a conjugative pKPC-CAV1312 plasmid of 320,480 bp, on a novel composite transposon (12,907 bp). FosA3 transposon shared similarities with other fosA3-harboring pKPC-CAV1312 plasmids among Citrobacter spp. We report the first case of FosA3 production in clinical carbapenemase-producing C. freundii ST116. The incidence of FosA3 enzymes is increasing among Enterobacterales, affecting even low-virulence pathogens, as C. freundii.

• MeSH
• antibakteriální látky * farmakologie   MeSH
• bakteriální proteiny * genetika   metabolismus   MeSH
• beta-laktamasy * genetika   metabolismus   MeSH
• Citrobacter freundii * genetika   enzymologie   účinky léků   MeSH
• enterobakteriální infekce * mikrobiologie   MeSH
• fosfomycin * farmakologie   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• mnohočetná bakteriální léková rezistence genetika   MeSH
• plazmidy genetika   MeSH
• sekvenování celého genomu MeSH
• transpozibilní elementy DNA MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Itálie MeSH