BACKGROUND: VIM metallo-β-lactamases are enzymes characterized by the ability to hydrolyze all β-lactams. Usually, bla VIM-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 bla VIM 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 bla VIM-1 allele while six isolates carried the bla VIM-4 gene. The most prevalent was the In110 integron (n = 24) and two isolates carried the In4873 class 1 integron. The bla VIM-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 bla VIM 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 bla VIM-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.

Biomedical Center Faculty of Medicine Charles University Pilsen Czechia

Department of Medical Microbiology 3rd Faculty of Medicine Charles University Prague Czechia

Department of Microbiology Faculty of Medicine University Hospital in Pilsen Charles University Pilsen Czechia

Department of Microbiology University Hospital of Larissa Larissa Greece

National Reference Laboratory for Antibiotics National Institute of Public Health Prague Czechia

Zobrazit více v PubMed

Bankevich A., Nurk S., Antipov D., Gurevich A. A., Dvorkin M., Kulikov A. S., et al. . (2012). SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J. Comput. Biol. 19, 455–477. doi: 10.1089/cmb.2012.0021, PMID: PubMed DOI PMC

Bitar I., Papagiannitsis C. C., Kraftova L., Chudejova K., Mattioni Marchetti V., Hrabak J. (2020). Detection of five mcr-9-carrying Enterobacterales isolates in four Czech hospitals. mSphere 5:e01008-20. doi: 10.1128/mSphere.01008-20 PubMed DOI PMC

Bolger A. M., Lohse M., Usadel B. (2014). Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30, 2114–2120. doi: 10.1093/bioinformatics/btu170, PMID: PubMed DOI PMC

Colinon C., Miriagou V., Carattoli A., Luzzaro F., Rossolini G. M. (2007). Characterization of the IncA/C plasmid pCC416 encoding VIM-4 and CMY-4 beta-lactamases. J. Antimicrob. Chemother. 60, 258–262. doi: 10.1093/jac/dkm171, PMID: PubMed DOI

Ellington M. J., Kistler J., Livermore D. M., Woodford N. (2007). Multiplex PCR for rapid detection of genes encoding acquired metallo-beta-lactamases. J. Antimicrob. Chemother. 59, 321–322. doi: 10.1093/jac/dkl481, PMID: PubMed DOI

European Committee on Antimicrobial Susceptibility Testing (EUCAST). (2012). EUCAST guidelines for detection of resistance mechanism and specific resistances of clinical and/or epidemiological importance. European committee on antimicrobial susceptibility testing. Växjö, Sweden: EUCAST.

Garcia-Fernandez A., Carattoli A. (2010). Plasmid double locus sequence typing for IncHI2 plasmids, a subtyping scheme for the characterization of IncHI2 plasmids carrying extended-spectrum beta-lactamase and quinolone resistance genes. J. Antimicrob. Chemother. 65, 1155–1161. doi: 10.1093/jac/dkq101, PMID: PubMed DOI

Garcia-Fernandez A., Villa L., Moodley A., Hasman H., Miriagou V., Guardabassi L., et al. . (2011). Multilocus sequence typing of IncN plasmids. J. Antimicrob. Chemother. 66, 1987–1991. doi: 10.1093/jac/dkr225, PMID: PubMed DOI

Halova D., Papousek I., Jamborova I., Masarikova M., Cizek A., Janecko N., et al. . (2014). Plasmid-mediated quinolone resistance genes in Enterobacteriaceae from American crows: high prevalence of bacteria with variable qnrB genes. Antimicrob. Agents Chemother. 58, 1257–1258. doi: 10.1128/AAC.01849-13, PMID: PubMed DOI PMC

Kraftova L., Finianos M., Studentova V., Chudejova K., Jakubu V., Zemlickova H., et al. . (2021). Evidence of an epidemic spread of KPC-producing Enterobacterales in Czech hospitals. Sci. Rep. 11:15732. doi: 10.1038/s41598-021-95285-z, PMID: PubMed DOI PMC

Laraki N., Galleni M., Thamm I., Riccio M. L., Amicosante G., Frere J. M., et al. . (1999). Structure of In31, a blaIMP-containing Pseudomonas aeruginosa integron phyletically related to In5, which carries an unusual array of gene cassettes. Antimicrob. Agents Chemother. 43, 890–901. doi: 10.1128/AAC.43.4.890, PMID: PubMed DOI PMC

Lombardi G., Luzzaro F., Docquier J. D., Riccio M. L., Perilli M., Coli A., et al. . (2002). Nosocomial infections caused by multidrug-resistant isolates of pseudomonas putida producing VIM-1 metallo-beta-lactamase. J. Clin. Microbiol. 40, 4051–4055. doi: 10.1128/JCM.40.11.4051-4055.2002, PMID: PubMed DOI PMC

Marchetti V. M., Bitar I., Sarti M., Fogato E., Scaltriti E., Bracchi C., et al. . (2021). Genomic characterization of VIM and MCR co-producers: the first two clinical cases, in Italy. Diagnostics (Basel) 11:0079. doi: 10.3390/diagnostics11010079, PMID: PubMed DOI PMC

Mazel D. (2006). Integrons: agents of bacterial evolution. Nat. Rev. Microbiol. 4, 608–620. doi: 10.1038/nrmicro1462, PMID: PubMed DOI

Mazel D., Davies J. (1999). Antibiotic resistance in microbes. Cell. Mol. Life Sci. 56, 742–754. doi: 10.1007/s000180050021 PubMed DOI PMC

Miriagou V., Papagiannitsis C. C., Kotsakis S. D., Loli A., Tzelepi E., Legakis N. J., et al. . (2010). Sequence of pNL194, a 79.3-kilobase IncN plasmid carrying the blaVIM-1 metallo-beta-lactamase gene in Klebsiella pneumoniae. Antimicrob. Agents Chemother. 54, 4497–4502. doi: 10.1128/AAC.00665-10, PMID: PubMed DOI PMC

Miriagou V., Tzelepi E., Gianneli D., Tzouvelekis L. S. (2003). Escherichia coli with a self-transferable, multiresistant plasmid coding for metallo-beta-lactamase VIM-1. Antimicrob. Agents Chemother. 47, 395–397. doi: 10.1128/AAC.47.1.395-397.2003, PMID: PubMed DOI PMC

Moura A., Soares M., Pereira C., Leitao N., Henriques I., Correia A. (2009). INTEGRALL: a database and search engine for integrons, integrases and gene cassettes. Bioinformatics 25, 1096–1098. doi: 10.1093/bioinformatics/btp105, PMID: PubMed DOI

Naas T., Cuzon G., Villegas M. V., Lartigue M. F., Quinn J. P., Nordmann P. (2008). Genetic structures at the origin of acquisition of the beta-lactamase Bla KPC gene. Antimicrob. Agents Chemother. 52, 1257–1263. doi: 10.1128/AAC.01451-07, PMID: PubMed DOI PMC

Papagiannitsis C. C., Dolejska M., Izdebski R., Giakkoupi P., Skalova A., Chudejova K., et al. . (2016). Characterisation of IncA/C2 plasmids carrying an In416-like integron with the blaVIM-19 gene from Klebsiella pneumoniae ST383 of Greek origin. Int. J. Antimicrob. Agents 47, 158–162. doi: 10.1016/j.ijantimicag.2015.12.001, PMID: PubMed DOI

Papagiannitsis C. C., Paskova V., Chudejova K., Medvecky M., Bitar I., Jakubu V., et al. . (2018). Characterization of pEncl-30969cz, a novel ColE1-like plasmid encoding VIM-1 carbapenemase, from an Enterobacter cloacae sequence type 92 isolate. Diagn. Microbiol. Infect. Dis. 91, 191–193. doi: 10.1016/j.diagmicrobio.2018.01.024, PMID: PubMed DOI

Papagiannitsis C. C., Studentova V., Ruzicka F., Tejkalova R., Hrabak J. (2013). Molecular characterization of metallo-beta-lactamase-producing Pseudomonas aeruginosa in a Czech hospital (2009-2011). J. Med. Microbiol. 62, 945–947. doi: 10.1099/jmm.0.056119-0, PMID: PubMed DOI

Papousek I., Papagiannitsis C. C., Medvecky M., Hrabak J., Dolejska M. (2017). Complete nucleotide sequences of two VIM-1-encoding plasmids from Klebsiella pneumoniae and Leclercia adecarboxylata isolates of Czech origin. Antimicrob. Agents Chemother. 61:2648. doi: 10.1128/AAC.02648-16, PMID: PubMed DOI PMC

Partridge S. R., Hall R. M. (2003). The IS1111 family members IS4321 and IS5075 have subterminal inverted repeats and target the terminal inverted repeats of Tn21 family transposons. J. Bacteriol. 185, 6371–6384. doi: 10.1128/JB.185.21.6371-6384.2003, PMID: PubMed DOI PMC

Poirel L., Heritier C., Tolun V., Nordmann P. (2004). Emergence of oxacillinase-mediated resistance to imipenem in Klebsiella pneumoniae. Antimicrob. Agents Chemother. 48, 15–22. doi: 10.1128/AAC.48.1.15-22.2004, PMID: PubMed DOI PMC

Ren Y., Ren Y., Zhou Z., Guo X., Li Y., Feng L., et al. . (2010). Complete genome sequence of Enterobacter cloacae subsp. cloacae type strain ATCC 13047. J. Bacteriol. 192, 2463–2464. doi: 10.1128/JB.00067-10, PMID: PubMed DOI PMC

Rotova V., Papagiannitsis C. C., Skalova A., Chudejova K., Hrabak J. (2017). Comparison of imipenem and meropenem antibiotics for the MALDI-TOF MS detection of carbapenemase activity. J. Microbiol. Methods 137, 30–33. doi: 10.1016/j.mimet.2017.04.003, PMID: PubMed DOI

Sheppard A. E., Stoesser N., Wilson D. J., Sebra R., Kasarskis A., Anson L. W., et al. . (2016). Nested Russian doll-like genetic mobility drives rapid dissemination of the Carbapenem resistance gene blaKPC. Antimicrob. Agents Chemother. 60, 3767–3778. doi: 10.1128/AAC.00464-16, PMID: PubMed DOI PMC

Tato M., Coque T. M., Baquero F., Canton R. (2010). Dispersal of carbapenemase blaVIM-1 gene associated with different Tn402 variants, mercury transposons, and conjugative plasmids in Enterobacteriaceae and Pseudomonas aeruginosa. Antimicrob. Agents Chemother. 54, 320–327. doi: 10.1128/AAC.00783-09, PMID: PubMed DOI PMC

Yong D., Toleman M. A., Giske C. G., Cho H. S., Sundman K., Lee K., et al. . (2009). Characterization of a new metallo-beta-lactamase gene, Bla(NDM-1), and a novel erythromycin esterase gene carried on a unique genetic structure in Klebsiella pneumoniae sequence type 14 from India. Antimicrob. Agents Chemother. 53, 5046–5054. doi: 10.1128/AAC.00774-09, PMID: PubMed DOI PMC

Genomic characterization of ST38 NDM-5-producing Escherichia coli isolates from an outbreak in the Czech Republic

Polyclonal Spread of Fosfomycin Resistance among Carbapenemase-Producing Members of the Enterobacterales in the Czech Republic