The aim of this study was to characterize four Enterobacterales co-producing NDM- and OXA-48-like carbapenemases from Czech patients with travel history or/and previous hospitalization abroad. Klebsiella pneumoniae isolates belonged to "high risk" clones ST147, ST11, and ST15, while the Escherichia coli isolate was assigned to ST167. All isolates expressed resistance against most β-lactams, including carbapenems, while retaining susceptibility to colistin. Furthermore, analysis of WGS data showed that all four isolates co-produced OXA-48- and NDM-type carbapenemases, in different combinations (Kpn47733: bla NDM- 5 + bla OXA- 181; Kpn50595: bla NDM- 1 + bla OXA- 181; Kpn51015: bla NDM- 1 + bla OXA- 244; Eco52418: bla NDM- 5 + bla OXA- 244). In Kpn51015, the bla OXA- 244 was found on plasmid p51015_OXA-244, while the respective gene was localized in the chromosomal contig of E. coli Eco52418. On the other hand, bla OXA- 181 was identified on a ColKP3 plasmid in isolate Kpn47733, while a bla OXA- 181-carrying plasmid being an IncX3-ColKP3 fusion was identified in Kpn50595. The bla NDM- 1 gene was found on two different plasmids, p51015_NDM-1 belonging to a novel IncH plasmid group and p51015_NDM-1 being an IncF K 1-FIB replicon. Furthermore, the bla NDM- 5 was found in two IncFII plasmids exhibiting limited nucleotide similarity to each other. In both plasmids, the genetic environment of bla NDM- 5 was identical. Finally, in all four carbapenemase-producing isolates, a diverse number of additional replicons, some of these associated with important resistance determinants, like bla CTX-M- 15, arr-2 and ermB, were identified. In conclusion, this study reports the first description of OXA-244-producing Enterobacterales isolated from Czech hospitals. Additionally, our findings indicated the genetic plurality involved in the acquisition and dissemination of determinants encoding OXA/NDM carbapenemases.

Biomedical Center Faculty of Medicine Charles University Pilsen Czechia

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

Department of Microbiology University Hospital of Larissa Larissa Greece

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Ahmad N., Ali S. M., Khan A. U. (2019). Molecular characterization of novel sequence type of carbapenem-resistant New Delhi metallo-β-lactamase-1-producing Klebsiella pneumoniae in the neonatal intensive care unit of an Indian hospital. Int. J. Antimicrob. Agents 53 525–529. 10.1016/j.ijantimicag.2018.12.005 PubMed DOI

Alcock B. P., Raphenya A. R., Lau T. T. Y., Tsang K. K., Bouchard M., Edalatmand A., et al. (2020). CARD 2020: antibiotic resistome surveillance with the comprehensive antibiotic resistance database. Nucleic Acids Res. 48 D517–D525. 10.1093/nar/gkz935 PubMed DOI PMC

Alikhan N. F., Petty N. K., Ben Zakour N. L., Beatson S. A. (2011). BLAST ring image generator (BRIG): simple prokaryote genome comparisons. BMC Genomics 12:402. 10.1186/1471-2164-12-402 PubMed DOI PMC

Allyn J., Coolen-Allou N., de Parseval B., Galas T., Belmonte O., Allou N., et al. (2018). Medical evacuation from abroad of critically ill patients: a case report and ethical issues. Medicine (Baltimore) 97:e12516. 10.1097/MD.0000000000012516 PubMed DOI PMC

Baek J. Y., Cho S. Y., Kim S. H., Kang C. I., Peck K. R., Song J. H., et al. (2019). Plasmid analysis of Escherichia coli isolates from South Korea co-producing NDM-5 and OXA-181 carbapenemases. Plasmid 104:102417. 10.1016/j.plasmid.2019.102417 PubMed DOI

Bakthavatchalam Y. D., Anandan S., Veeraraghavan B. (2016). Laboratory detection and clinical implication of oxacillinase-48 like carbapenemase: the hidden threat. J. Glob. Infect. Dis. 8 41–50. 10.4103/0974-777X.176149 PubMed DOI PMC

Balm M. N., La M. V., Krishnan P., Jureen R., Lin R. T., Teo J. W. (2013). Emergence of Klebsiella pneumoniae co-producing NDM-type and OXA-181 carbapenemases. Clin. Microbiol. Infect. 19 E421–E423. 10.1111/1469-0691.12247 PubMed DOI

Bassetti M., Peghin M., Vena A., Giacobbe D. R. (2019). Treatment of infections due to MDR gram-negative bacteria. Front. Med. 6:74. 10.3389/fmed.2019.00074 PubMed DOI PMC

Carattoli A., Zankari E., García-Fernández A., Voldby Larsen M., Lund O., Villa L., et al. (2014). In silico detection and typing of plasmids using PlasmidFinder and plasmid multilocus sequence typing. Antimicrob. Agents Chemother. 7 3895–3903. PubMed PMC

Castanheira M., Deshpande L. M., Mathai D., Bell J. M., Jones R. N., Mendes R. E. (2011). Early dissemination of NDM-1-and OXA-181-producing Enterobacteriaceae in Indian hospitals: report from the SENTRY antimicrobial surveillance program, 2006-2007. Antimicrob. Agents Chemother. 55 1274–1278. PubMed PMC

Contreras D. A., Fitzwater S. P., Nanayakkara D. D., Schaenman J., Aldrovandi G. M., Garner O. B., et al. (2020). Coinfections of two strains of NDM-1-and OXA-232-coproducing Klebsiella pneumoniae in a kidney transplant patient. Antimicrob. Agents Chemother. 64:e00948-19. 10.1128/AAC.00948-19 PubMed DOI PMC

Doi Y., O’Hara J. A., Lando J. F., Querry A. M., Townsend B. M., Pasculle A. W., et al. (2014). Co-production of NDM-1 and OXA-232 by Klebsiella pneumoniae. Emerg. Infect. Dis. 20:163. 10.3201/eid2001.130904 PubMed DOI PMC

Doi Y., Potoski B. A., Adams-Haduch J. M., Sidjabat H. E., Pasculle A. W., Paterson D. L. (2008). Simple disk-based method for detection of Klebsiella pneumoniae carbapenemase-type -lactamase by use of a boronic acid compound. J. Clin. Microbiol. 46 4083–4086. PubMed PMC

Fursova N. K., Astashkin E. I., Knyazeva A. I., Kartsev N. N., Leonova E. S., Ershova O. N., et al. (2015). The spread of blaOXA–48 and blaOXA–244 carbapenemase genes among Klebsiella pneumoniae, Proteus mirabilis and Enterobacter spp. isolated in Moscow, Russia. Ann. Clin. Microbiol. Antimicrob. 14:46. 10.1186/s12941-015-0108-y PubMed DOI PMC

Glupczynski Y., Huang T. D., Bouchahrouf W., Rezende de Castro R., Bauraing C., Gérard M., et al. (2012). Rapid emergence and spread of OXA-48-producing carbapenem-resistant Enterobacteriaceae isolates in Belgian hospitals. Int. J. Antimicrob. Agents 39 168–172. PubMed

Gupta V., Ye G., Olesky M., Lawrence K., Murray J., Yu K. (2019). Trends in resistant Enterobacteriaceae and Acinetobacter species in hospitalized patients in the United States: 2013–2017. BMC Infect. Dis. 19:742. 10.1186/s12879-019-4387-3 PubMed DOI PMC

Hammerum A. M., Porsbo L. J., Hansen F., Roer L., Kaya H., Henius A., et al. (2020). Surveillance of OXA-244-producing Escherichia coli and epidemiologic investigation of cases, Denmark, January 2016 to August 2019. Euro Surveill. 25:1900742 10.2807/1560-7917.ES.2020.25.18.1900742 PubMed DOI PMC

Hao Y., Shao C., Geng X., Bai Y., Jin Y., Lu Z. (2019). Genotypic and phenotypic characterization of clinical Escherichia coli sequence type 405 carrying IncN2 plasmid harboring blaNDM–1. Front. Microbiol. 10:788. 10.3389/fmicb.2019.00788 PubMed DOI PMC

Holman A. M., Allyn J., Miltgen G., Lugagne N., Traversier N., Picot S., et al. (2017). Surveillance of carbapenemase-producing Enterobacteriaceae in the Indian ocean region between January 2010 and December 2015. Med. Mal. Infect. 47 333–339. 10.1016/j.medmal.2017.04.007 PubMed DOI

Hornsey M., Phee L., Wareham D. W. (2011). A novel variant, NDM-5, of the New Delhi metallo-β-lactamase in a multidrug-resistant Escherichia coli ST648 isolate recovered from a patient in the United Kingdom. Antimicrob. Agents Chemother. 55 5952–5954. PubMed PMC

Hoyos-Mallecot Y., Naas T., Bonnin R. A., Patino R., Glaser P., Fortineau N., et al. (2017). OXA-244-producing Escherichia coli isolates, a challenge for clinical microbiology laboratories. Antimicrob. Agents Chemother. 61:e00818-17. 10.1128/AAC.00818-17 PubMed DOI PMC

Hrabak J., Stolbova M., Studentova V., Fridrichova M., Chudackova E., Zemlickova H. (2012). NDM-1 producing Acinetobacter baumannii isolated from a patient repatriated to the Czech Republic from Egypt, July 2011. Euro Surveill. 17:20085. PubMed

Kaase M., Nordmann P., Wichelhaus T. A., Gatermann S. G., Bonnin R. A., Poirel L. (2011). NDM-2 carbapenemase in Acinetobacter baumannii from Egypt. J. Antimicrob. Chemother. 66 1260–1262. PubMed

Krishnaraju M., Kamatchi C., Jha A. K., Devasena N., Vennila R., Sumathi G., et al. (2015). Complete sequencing of an IncX3 plasmid carrying blaNDM–5 allele reveals an early stage in the dissemination of the blaNDM gene. Indian J. Med. Microbiol. 33 30–38. 10.4103/0255-0857.148373 PubMed DOI

Larsen M. V., Cosentino S., Rasmussen S., Friis C., Hasman H., Marvig R. L., et al. (2012). Multilocus sequence typing of total genome sequenced bacteria. J. Clin. Micobiol. 50 1355–1361. PubMed PMC

Lascols C., Peirano G., Hackel M., Laupland K. B., Pitout J. D. (2013). Surveillance and molecular epidemiology of Klebsiella pneumoniae isolates that produce carbapenemases: first report of OXA-48-like enzymes in North America. Antimicrob. Agents Chemother. 57 130–136. 10.1128/AAC.01686-12 PubMed DOI PMC

Lee K., Lim Y. S., Yong D., Yum J. H., Chong Y. (2003). Evaluation of the hodge test and the imipenem-EDTA double-disk synergy test for differentiating Metallo-β-Lactamase-producing isolates of Pseudomonas spp. and Acinetobacter spp. J. Clin. Microbiol. 41 4623–4629. PubMed PMC

Mairi A., Pantel A., Sotto A., Lavigne J. P., Touati A. (2018). OXA-48-like carbapenemases producing Enterobacteriaceae in different niches. Eur. J. Clin. Microbiol. Infect. Dis. 37 587–604. PubMed

Mani Y., Mansour W., Mammeri H., Denamur E., Saras E., Boujâafar N., et al. (2017). KPC-3-producing ST167 Escherichia coli from mussels bought at a retail market in Tunisia. J. Antimicrob. Chemother. 72 2403–2404. PubMed

Marchetti V. M., Bitar I., Mercato A., Nucleo E., Bonomini A., Pedroni P., et al. (2019). Complete nucleotide sequence of plasmids of two Escherichia coli strains carrying blaNDM–5 and blaOXA–181 From the Same Patient. Front. Microbiol. 10:3095. 10.3389/fmicb.2019.03095 PubMed DOI PMC

McConville T. H., Sullivan S. B., Gomez-Simmonds A., Whittier S., Uhlemann A. C. (2017). Carbapenem-resistant Enterobacteriaceae colonization (CRE) and subsequent risk of infection and 90-day mortality in critically ill patients, an observational study. PLoS One 12:10. 10.1371/journal.pone.0186195 PubMed DOI PMC

Nordmann P., Boulanger A. E., Poirel L. (2012). NDM-4 metallo-β-lactamase with increased carbapenemase activity from Escherichia coli. Antimicrob. Agents Chemother. 56 2184–2186. PubMed PMC

Nordmann P., Poirel L. (2014). The difficult-to-control spread of carbapenemase producers among Enterobacteriaceae worldwide. Clin. Microbiol. Infect. 20 821–830. PubMed

Nordmann P., Poirel L., Walsh T. R., Livermore D. M. (2011). The emerging NDM carbapenemases. Trends Microbiol. 19 588–595. 10.1016/j.tim.2011.09.005 PubMed DOI

Nucleo E., Marchetti V. M., Mercato A., Quatela M., Villa L., Migliavacca R. (2020). OXA-48 and NDM-1 Klebsiella pneumoniae of Sequence Type 101 from blood in a patient with travel history abroad. Italy. New Microbiol. 43 41–43. PubMed

Oteo J., Hernández J. M., Espasa M., Fleites A., Sáez D., Bautista V., et al. (2013). Emergence of OXA-48-producing Klebsiella pneumoniae and the novel carbapenemases OXA-244 and OXA-245 in Spain. J. Antimicrob. Chemother. 68 317–321. PubMed

Otlu B., Yakupoğulları Y., Gürsoy N. C., Duman Y., Bayındır Y., Tekerekoğlu M. S., et al. (2018). Co-production of OXA-48 and NDM-1 carbapenemases in Providencia rettgeri: the first report. Mikrobiyol. Bul. 52 300–307. 10.5578/mb.67153 PubMed DOI

Papagiannitsis C. C., Izdebski R., Baraniak A., Fiett J., Herda M., Hrabák J., et al. (2015). Survey of metallo-β-lactamase-producing Enterobacteriaceae colonizing patients in European ICUs and rehabilitation units, 2008-11. J. Antimicrob. Chemother. 70 1981–1988. PubMed

Paskova V., Medvecky M., Skalova A., Chudejova K., Bitar I., Jakubu V., et al. (2018). Characterization of NDM-Encoding plasmids from Enterobacteriaceae recovered from czech hospitals. Front. Microbiol. 9:1549. 10.3389/fmicb.2018.01549 PubMed DOI PMC

Pitout J. D. D., Peirano G., Kock M. M., Strydom K. A., Matsumura Y. (2019). The global ascendency of OXA-48-type carbapenemases. Clin. Microbiol. Rev. 33:e00102-19. 10.1128/CMR.00102-19 PubMed DOI PMC

Poirel L., Héritier C., Tolün V., Nordmann P. (2004). Emergence of oxacillinase-mediated resistance to imipenem in Klebsiella pneumoniae. Antimicrob. Agents Chemother. 48 15–22. PubMed PMC

Poirel L., Lascols C., Bernabeu S., Nordmann P. (2012). NDM-1-producing Klebsiella pneumoniae in Mauritius. Antimicrob. Agents Chemother. 56 598–599. 10.1128/AAC.05639-11 PubMed DOI PMC

Potron A., Nordmann P., Lafeuille E., Al Maskari Z., Al Rashdi F., Poirel L. (2011). Characterization of OXA-181, a carbapenem-hydrolyzing class D beta-lactamase from Klebsiella pneumoniae. Antimicrob. Agents Chemother. 55 4896–4899. PubMed PMC

Potron A., Poirel L., Dortet L., Nordmann P. (2016). Characterisation of OXA-244, a chromosomally-encoded OXA-48-like β-lactamase from Escherichia coli. Int. J. Antimicrob. Agents 47 102–103. PubMed

Principe L., Mauri C., Conte V., Pini B., Giani T., Rossolini G. M., et al. (2016). First report of NDM-1-producing Klebsiella pneumoniae imported from Africa to Italy: evidence of the need for continuous surveillance. J. Glob. Antimicrob. Resist. 8 23–27. 10.1016/j.jgar.2016.10.004 PubMed DOI

Roberts L. W., Harris P. N., Forde B. M., Zakour N. L. B., Catchpoole E., Stanton-Cook M., et al. (2020). Integrating multiple genomic technologies to investigate an outbreak of carbapenemase-producing Enterobacter hormaechei. Nat. Commun. 11:466. 10.1038/s41467-019-14139-5 PubMed DOI PMC

Rogers B. A., Sidjabat H. E., Silvey A., Anderson T. L., Perera S., Li J., et al. (2013). Treatment options for New Delhi metallo-beta-lactamase-harboring Enterobacteriaceae. Microb. Drug Resist. 19 100–103. PubMed

Rojas L. J., Hujer A. M., Rudin S. D., Wright M. S., Domitrovic T. N., Marshall S. H., et al. (2017). NDM-5 and OXA-181 beta-lactamases, a significant threat continues to spread in the Americas. Antimicrob. Agents Chemother. 61:e00454-17. 10.1128/AAC.00454-17 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. PubMed

Sánchez-Benito R., Iglesias M. R., Quijada N. M., Campos M. J., Ugarte-Ruiz M., Hernández M., et al. (2017). Escherichia coli ST167 carrying plasmid mobilisable mcr-1 and blaCTX–M–15 resistance determinants isolated from a human respiratory infection. Int. J. Antimicrob. Agents 50 285–286. PubMed

Siguier P., Perochon J., Lestrade L., Mahillon J., Chandler M. (2006). ISfinder: the reference centre for bacterial insertion sequences. Nucleic Acids Res. 34 D32–D36. PubMed PMC

Skalova A., Chudejova K., Rotova V., Medvecky M., Studentova V., Chudackova E., et al. (2017). Molecular Characterization of OXA-48-like-producing Enterobacteriaceae in the Czech Republic and evidence for horizontal transfer of pOXA-48-like plasmids. Antimicrob. Agents Chemother. 61:e01889-16. PubMed PMC

Soliman A. M., Ramadan H., Sadek M., Nariya H., Shimamoto T., Hiott L. M., et al. (2020a). Draft genome sequence of a blaNDM–1- and blaOXA–244-carrying multidrug-resistant Escherichia coli D-ST69 clinical isolate from Egypt. J. Glob. Antimicrob. Resist. 22 832–834. 10.1016/j.jgar.2020.07.015 PubMed DOI

Soliman A. M., Zarad H. O., Nariya H., Shimamoto T., Shimamoto T. (2020b). Genetic analysis of carbapenemase-producing Gram-negative bacteria isolated from a university teaching hospital in Egypt. Infect. Genet. Evol. 77:104065. 10.1016/j.meegid.2019.104065 PubMed DOI

Sun L., Xu J., He F. (2018). Draft genome sequence of an NDM-5, CTX-M-15 and OXA-1 co-producing Escherichia coli ST167 clinical strain isolated from a urine sample. J. Glob. Antimicrob. Resist. 14 284–286. PubMed

Tada T., Miyoshi-Akiyama T., Dahal R. K., Sah M. K., Ohara H., Kirikae T., et al. (2013). NDM-8 metallo-β-lactamase in a multidrug-resistant Escherichia coli strain isolated in Nepal. Antimicrob. Agents Chemother. 57 2394–2396. PubMed PMC

Tafoukt R., Touati A., Leangapichart T., Bakour S., Rolain J. M. (2017). Characterization of OXA-48-like-producing Enterobacteriaceae isolated from river water in Algeria. Water Res. 120 185–189. 10.1016/j.watres.2017.04.073 PubMed DOI

van Hattem J. M., Arcilla M. S., Bootsma M. C., van Genderen P. J., Goorhuis A., Grobusch M. P., et al. (2016). Prolonged carriage and potential onward transmission of carbapenemase-producing Enterobacteriaceae in Dutch travelers. Future Microbiol. 11 857–864. PubMed

Villa L., Poirel L., Nordmann P., Carta C., Carattoli A. (2012). Complete sequencing of an IncH plasmid carrying the blaNDM–1, blaCTX–M–15 and qnrB1 genes. J. Antimicrob. Chemother. 67 1645–1650. PubMed

Wang X., Li H., Zhao C., Chen H., Liu J., Wang Z., et al. (2014). Novel NDM-9 metallo-β-lactamase identified from a ST107 Klebsiella pneumoniae strain isolated in China. Inter. J. Antimicrob. Agents 44:90. 10.1016/j.ijantimicag.2014.04.010 PubMed DOI

Williamson D. A., Sidjabat H. E., Freeman J. T., Roberts S. A., Silvey A., Woodhouse R., et al. (2012). Identification and molecular characterisation of New Delhi metallo-β-lactamase-1 (NDM-1)- and NDM-6-producing Enterobacteriaceae from New Zealand hospitals. Inter. J. Antimicrob. Agents 39 529–533. PubMed

Woodford N., Turton J. F., Livermore D. M. (2011). Multiresistant Gram-negative bacteria: the role of high-risk clones in the dissemination of antibiotic resistance. FEMS Microbiol. Rev. 35 736–755. PubMed

Wu W., Feng Y., Tang G., Qiao F., McNally A., Zong Z. (2019). NDM metallo-β-lactamases and their bacterial producers in health care settings. Clin. Microbiol. Rev. 32:e00115-18. 10.1128/CMR.00115-18 PubMed DOI PMC

Xu L., Wang P., Cheng J., Qin S., Xie W. (2019). Characterization of a novel blaNDM–5-harboring IncFII plasmid and an mcr-1-bearing IncI2 plasmid in a single Escherichia coli ST167 clinical isolate. Infect. Drug Resist. 12 511–519. PubMed PMC

Ye Y., Xu L., Han Y., Chen Z., Liu C., Ming L. (2018). Mechanism for carbapenem resistance of clinical Enterobacteriaceae isolates. Exp. Ther. Med. 15 1143–1149. PubMed PMC

Yong D., Toleman M. A., Giske C. G., Cho H. S., Sundman K., Lee K., et al. (2009). Characterization of a new metallo-β-lactamase gene, blaNDM–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. PubMed PMC

Zankari E., Henrik H., Salvatore C., Martin V., Simon R., Ole L., et al. (2012). Identification of acquired antimicrobial resistance genes. J. Antimicrob. Chemother. 11 2640–2644. PubMed PMC

Zingali T., Chapman T. A., Webster J., Chowdhury P. R., Djordjevic S. P. (2020). Genomic characterisation of a multiple drug resistant IncHI2 ST4 plasmid in Escherichia coli ST744 in Australia. Microorganisms 8:896. 10.3390/microorganisms8060896 PubMed DOI PMC

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