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Carbapenemase-Producing Gram-Negative in Andalusia, Spain, 2014–2018 Inmaculada López-Hernández, Mercedes Delgado-Valverde, Felipe Fernández-Cuenca, Lorena López-Cerero, Jesús Machuca, Álvaro Pascual

The emergence and spread of carbapenemase-producing (https://www.iavante.es/es/programa-pirasoa) gram-negative bacteria is a major public health concern. program was designed to prevent and control We used data collected from microbiology laboratories as healthcare-associated infections and promote the part of the PIRASOA program during 2014–2018 to study appropriate use of antimicrobials, covering the en- the epidemiology of carbapenemase-producing bacteria tire area of the Andalusia public healthcare system, in Andalusia, Spain. Our findings highlight the importance which includes 34 hospitals in 8 provinces. Data col- of ongoing surveillance and epidemiologic studies for lected through PIRASOA during 2014–2018 provid- these bacteria. ed an opportunity for us to study and understand the epidemiology of carbapenemase-producing bac- here are 3 common carbapenemase classes: class teria in Andalusia. TA includes pneumoniae carbapenemase (KPC); class B is metallo-β-lactamases (MBL), includ- The Study ing Verona integron-encoded metallo-β-lactamase During January 2014–December 2018, microbiology (VIM), New Delhi metallo-β-lactamase (NDM) and laboratories in the hospitals included in the PIRA- imipenemase (IMP); and class D includes oxacillin- SOA program voluntarily submitted CPGNB de- ases (OXA), including OXA-48–like carbapenemases. tected according to EUCAST screening criteria (6) to In Spain, all 3 of these classes have been reported in the PIRASOA reference laboratory. We identified the recent studies (1–4). A multicenter study of Enterobac- isolates using MALDI-TOF Biotyper (Bruker Dalton- terales carbapenemase-producing gram-negative bac- ics, https://www.bruker.com). Antimicrobial sus- teria (CPGNB), isolated from urine specimens, found ceptibility testing was performed by microdilution a prevalence of 1.6%. OXA-48 was the most prevalent, (Microscan; Beckman Coulter, https://www.beck- followed by KPC and MBL (5). mancoulter.com) and interpreted according to EU- In Andalusia, a region in the south of Spain with CAST breakpoints (7). We determined the inhibition a population of ≈8.4 million people, The PIRASOA of carbapenemase activity by using inhibitors (dip- icolinic acid, phenyl-boronic acid, and cloxacillin). In Author affiliations: Laboratorio PIRASOA, Hospital Universitario addition, we tested all the isolates with the β-CARBA Virgen Macarena, Seville, Spain (I. Lopez-Hernandez, test (Bio-Rad, https://www.bio-rad.com) and, from M. Delgado-Valverde, F. Fernández-Cuenca, L. López-Cerero, January 2018, also with the NG-test CARBA 5 (NG J. Machuca, Á. Pascual); Instituto de Biomedicina de Sevilla IBIS, Biotech, https://ngbiotech.com). All isolates with Hospital Universitario Virgen Macarena, Consejo Superior de a positive or indeterminate result from phenotypic Investigaciones Científicas, Universidad de Sevilla, Seville screening were tested by PCR and sequencing (Mac- (I. López-Hernández, M. Delgado-Valverde, F. Fernández- rogen, https://dna.macrogen.com) (bla , bla , Cuenca, L. López-Cerero, J. Machuca, Á. Pascual); Red Española KPC OXA-48 bla , bla , bla , bla ) or by whole ge- de Investigación en Patología Infecciosa (REIPI RD16/0016), VIM IMP NDM OXA-23,24/40,58 nome sequencing on a MiSeq system (Illumina, Instituto de Salud Carlos III, Madrid, Spain (I. López-Hernández, https://www.illumina.com), which became avail- M. Delgado-Valverde, F. Fernández-Cuenca, L. López-Cerero, able in October 2017. We evaluated genetic related- J. Machuca, Á. Pascual); Universidad de Sevilla, Seville ness by pulsed-field gel electrophoresis (CHEF DR-II (Á. Pascual). system; Bio-Rad); isolates of <2 bands difference DOI: https://doi.org/10.3201/eid2609.191772 were assigned to the same cluster. We determined

2218 Emerging Infectious Diseases • www.cdc.gov/eid • Vol. 26, No. 9, September 2020 Carbapenemase-Producing Gram-Negative Bacteria multilocus sequence typing using PCR and sequenc- Over the course of the study period, we observed ing or whole-genome sequencing data (https://cge. an increase in MBL carbapenemases, with a note- cbs.dtu.dk/services/MLST). worthy increase in 2018 (Table 1; Figure 1). Klebsiella During the study period, we analyzed 2,005 was most prevalent MBL, comprising 53% (157/294), gram-negative isolates of which 1,243 (62%) were almost equally distributed between K. pneumoniae carbapenemase producers. Among those 1,243 iso- and K. oxytoca, followed by lates, clinical specimens (64%, 791/1,243) comprised (20%, 60/294) and (14%, 42/294). the predominant source, followed by surveillance The distribution of each MBL enzyme type varied (31%, 387/1,243) and environmental samples (5%, significantly. We found 39% (77/197) of VIM in K. 65/1,243). The most common species were K. pneu- oxytoca, 80% (45/56) of NDM in K. pneumoniae, and moniae (45%, 560/1,243), 73% (30/41) of IMP in P. aeruginosa. OXA-23 (49%, (34%, 425/1,243), and K. oxytoca (7%, 83/1,243). 208/425) was the most common carbapenemase pro- KPC-3 was the most common carbapenemase we ducer in A. baumannii, followed by OXA-58 (38%, found (249; 20%); 96% of the KPC-3 was detected in 162/425), and OXA-24/40 (13%, 55/425). OXA-23 K. pneumoniae belonging to clones sequence type (ST) and OXA 24/40 isolates belonged predominantly to 512 or ST258. In contrast, KPC-2, which comprised clone ST2. OXA-58 producers were detected in mul- only 1.8% (23), was found in multiple species. Among tiple clones. We summarize the distribution of car- OXA-48–like carbapenemases, which comprised 19% bapenemases among CPGNB in this study in Table 2. (254) of carbapenemases, 87% (221/254) were found Geographically, carbapenemase producers were in K. pneumoniae, 70% (155/221) of those in clones found in 32 hospitals distributed throughout 8 prov- ST11, ST15, ST392, or ST307. inces of the region, with KPC detected in 26 hospitals,

Table 1. Distribution of carbapenemase-producing gram-negative bacteria by year, Andalusia, Spain, 2014–2018 Year Carbapenemase Bacteria 2014 2015 2016 2017 2018 VIM 0 0 9 9 15 K. oxytoca 1 13 12 19 32 0 2 1 3 2 Enterobacter cloacae 0 4 4 10 15 0 1 3 2 2 C. amalonaticus 0 0 1 0 0 Pseudomonas aeruginosa 1 0 0 0 29 P. putida 0 0 0 3 3 Acinetobacter spp 0 0 0 1 0 IMP K. pneumoniae 0 0 0 0 2 E. coli 0 0 0 0 1 E. cloacae 0 0 0 2 5 C. freundii 0 1 0 0 0 P. aeruginosa 0 2 0 10 18 NDM K. pneumoniae 0 0 1 7 37 E. coli 0 0 1 1 3 E. cloacae 0 0 2 0 0 Acinetobacter spp 0 0 0 0 1 A. baumannii 0 0 0 2 1 KPC K. pneumoniae 35 71 61 54 38 K. oxytoca 0 0 0 0 2 E. coli 0 0 0 0 3 E. cloacae 0 0 2 3 2 C. freundii 0 0 0 1 0 OXA-48 K. pneumoniae 35 26 39 77 44 K. oxytoca 0 0 0 1 3 E. coli 6 3 0 1 3 E. cloacae 1 1 0 0 5 C. freundii 0 1 0 1 3 K. aerogenes 0 0 0 0 4 OXA-23 A. baumannii 38 30 19 67 54 OXA-58 A. baumannii 15 25 46 38 36 OXA-24/40 A. baumannii 1 33 12 7 1 IMI E. cloacae 0 0 0 0 1 IMP, imipenemase; KPC, Klebsiella pneumoniae carbapenemase; MBL, metallo-β-lactamases; NDM, New Delhi metallo-beta-lactamase; OXA, oxacillinases; VIM, Verona integron-encoded metallo-β-lactamase.

Emerging Infectious Diseases • www.cdc.gov/eid • Vol. 26, No. 9, September 2020 2219 DISPATCHES

Figure 1. Distribution of carbapenemase types recovered in Andalusia, 2014–2018. IMP, imipenemase; KPC, Klebsiella pneumoniae carbapenemase; MBL, metallo-β-lactamases; NDM, New Delhi metallo-beta- lactamase; OXA, oxacillinases; VIM, Verona integron-encoded metallo-β-lactamase.

followed by OXA-48–like in 23 and MBL in 18 (Ap- Conclusions pendix Figure). Sequence type was available for 1,181 Our results provide an overview of the molecular of 1,243 (95%) isolates. Some circulating high-risk epidemiology of CPGNB in Andalusia. The predomi- clones were detected in K. pneumoniae, E. cloacae, Esch- nance of K. pneumoniae as a producer of all classes of erichia coli, A. baumannii, and P. aeruginosa. The com- carbapenemases is noteworthy, reflecting its role in binations of species, carbapenemase variants, and se- disseminating carbapenemases in the region. KPC-3 quence types are listed in the Appendix Table. and OXA-48 were the carbapenemases most frequent- Six isolates (0.5%) produced 2 types of carbapene- ly detected. KPC-3 was almost exclusively associated mase: Five isolates combined VIM-1 with either OXA- with K. pneumoniae and importantly with high-risk 48 (K. oxytoca ST206 and E. cloacae ST114), OXA-245 clones ST512 and ST258 considered endemic in other (Citrobacter freundii ST63), IMP-8 (E. cloacae ST242), or European countries (8–10). Like KPC-3, OXA-48 was NDM-1 (C. freundii ST170). The remaining isolate was predominantly found in K. pneumoniae, but it began E. coli ST617 harboring NDM-5 and OXA-48. Accord- appearing in other species in 2018, which raises con- ing to pulsed-field gel electrophoresis, 765 of 1,243 cerns about the control of this carbapenemase in the isolates (62%) were included in 23 clusters submitted region. In addition, the high-risk clones of OXA-48– from 22 hospitals (Figure 2). producing K. pneumoniae, ST11, ST15, ST392, ST307,

Table 2. Distribution of carbapenemases by species, Andalusia, Spain, 2014–2018* Species Enterobacter Pseudomonas P. Acinetobacter A. K. p. K. o. E. coli cloacae C. f. C. a. aeruginosa putida spp. baumannii K. a. VIM-1 33 77 8 33 8 1 1 5 1 0 0 VIM-2 0 0 0 0 0 0 29 1 0 0 0 IMP-8 2 0 1 6 1 0 14 0 0 0 0 IMP-16 0 0 0 0 0 0 9 0 0 0 0 IMP-23 0 0 0 0 0 0 7 0 0 0 0 NDM-1 6 0 0 2 0 0 0 0 1 3 0 NDM-7 39 0 1 0 0 0 0 0 0 0 0 NDM-5 0 0 4 0 0 0 0 0 0 0 0 IMP-22 0 0 0 1 0 0 0 0 0 0 0 KPC-3 248 0 0 1 0 0 0 0 0 0 0 KPC-2 10 2 3 6 1 0 0 0 0 0 0 OXA-48 205 4 12 7 4 0 0 0 0 0 4 OXA-23 0 0 0 0 0 0 0 0 0 208 0 OXA-58 0 0 0 0 0 0 0 0 0 160 0 OXA-24/40 0 0 0 0 0 0 0 0 0 54 0 OXA-245 15 0 1 0 1 0 0 0 0 0 0 OXA-181 1 0 0 0 0 0 0 0 0 0 0 KPC-31 1 0 0 0 0 0 0 0 0 0 0 IMI-1 0 0 0 1 0 0 0 0 0 0 0 Total 560 83 30 57 15 1 60 6 2 425 4 *C. f., Citrobacter freundii; C. a., Citrobacter amalonaticus; E., Escherichia; K. a., ; K. o., K. oxytoca; K. p., Klebsiella pneumoniae. IMP, imipenemase; KPC, Klebsiella pneumoniae carbapenemase; MBL, metallo-β-lactamases; NDM, New Delhi metallo-beta-lactamase; OXA, oxacillinases; VIM, Verona integron-encoded metallo-β-lactamase.

2220 Emerging Infectious Diseases • www.cdc.gov/eid •Vol. 26, No. 9, September 2020 Carbapenemase-Producing Gram-Negative Bacteria

Figure 2. Characteristics of clusters of carbapenemase-producing gram negative bacteria, Andalusia, 2014–2018. Hospitals are identified by number: Almería ,1, 2, 5; Cádiz, 7, 8, 9; Córdoba, 6, 11, 12; Granada, 4, 13; Huelva, 20; Jaén, 10, 21; Málaga, 17, 18, 19; Sevilla, 3, 14, 15, 16. Ab, Acinetobacter baumannii; CTX-M, Cefotaximase-Munich; Eclo, Enterobacter cloacae; IMP, imipenemase; Kox, Klebsiella oxytoca; KPC, Klebsiella pneumoniae carbapenemase; Kpn, Klebsiella pneumoniae; MBL, metallo-β-lactamases; NDM, New Delhi metallo-beta-lactamase; OXA, oxacillinases; Pae, Pseudomonas aeruginosa; Pp, Pseudomonas putida; ST, sequence type; VIM, Verona integron-encoded metallo-β-lactamase. differed from those for KPC-3, showing that specific remarkable finding was the detection of A. baumannii clones have a role in the dissemination of specific car- ST85 harboring NDM-1 for the first time in Spain (14). bapenemases in this region. The increase of carbapenemases in E. coli is a finding We found a significant temporal change in the that could have epidemiologic significance becauseE. epidemiology of MBL with a clear upward trend as- coli spread in the community more readily than other sociated with several outbreaks in 2018. Findings species and could begin to establish carbapenemases from recent studies from Spain reflect the emergence in the community. It is also remarkable that 7 of 23 of MBL in different CPGNB species (4,11,12). VIM clusters were detected in the same hospital, reflecting was the MBL most frequently detected and, most of the burden of antimicrobial resistance in this hospital. the VIM-producing bacteria were detected as part of This issue remains serious, and additional efforts for clusters. IMP was almost exclusively found in P. aeru- infection control should be made. ginosa isolates, showing the importance of this species One limitation of this work is that, because sub- in the dissemination of this carbapenemase; it was as- mission of isolates is not mandatory, underreporting sociated with different clusters in a single hospital is likely. Nevertheless, we have evidence that most and each variant was associated with a specific clone: relevant isolates are routinely submitted. Further- IMP-8/ST348, IMP-23/ST175 and IMP-16/ST253. An more, this study reinforces the importance of sub- increase in NDM was observed in 2018 associated mitting all multidrug-resistant isolates to a reference with a large outbreak produced by K. pneumoniae laboratory because the information provided using ST11 NDM-7 in a single hospital. molecular techniques makes the rapid detection of In A. baumanni, OXA-23 and OXA-58 were the clusters and high-risk clones possible, which in turn, most common carbapenemases. ST2 was the pre- contributes to the success of infection prevention and dominant clone according to previous studies (13). A control programs.

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In summary, our study shows dissemination and carbapenemase-producing from urine establishment of CPGNB in Andalusia, including specimens obtained during the infection-carbapenem resistance evaluation surveillance trial (iCREST) in Spain. high-risk clones of different species. These findings Int J Antimicrob Agents. 2018;51:511–5. https://doi.org/ demonstrate the importance of continuing surveil- 10.1016/j.ijantimicag.2018.01.011 lance programs and epidemiologic studies to detect 6. EUCAST Guidelines for detection of resistance mechanisms and investigate the spread of carbapenemase-produc- and specific resistances of clinical and/or epidemiological importance, version 2.0 [cited 2 Jan 2018]. http://www. ing bacteria, particularly in Andalusia, which is a re- eucast.org/fileadmin/src/media/PDFs/EUCAST_files/ gion predisposed to the introduction of new lineages Resistance_mechanisms/EUCAST_detection_of_resistance_ because of tourism and migration pathways. mechanisms_170711.pdf 7. EUCAST. Breakpoint tables for interpretation of MICs and zone diameters, version 8.0, 2018 [cited 11 July 2017]. http://www. About the Author eucast.org/fileadmin/src/media/PDFs/EUCAST_files/ Dr. López-Hernández is a clinical microbiologist in the Breakpoint_tables/v_8.1_Breakpoint_Tables.pdf Unit of Infectious Diseases and Clinical Microbiology of 8. David S, Reuter S, Harris SR, Glasner C, Feltwell T, Argimon S, et al.; EuSCAPE Working Group; ESGEM the Virgen Macarena University Hospital, Seville, Spain. Study Group. Epidemic of carbapenem-resistant Klebsiella Her research interest is antimicrobial resistance pneumoniae in Europe is driven by nosocomial spread. Nat mechanisms in bacteria. Microbiol. 2019;4:1919–29. https://doi.org/10.1038/ s41564-019-0492-8 9. Ferrari C, Corbella M, Gaiarsa S, Comandatore F, Scaltriti E, References Bandi C, et al. Multiple Klebsiella pneumoniae KPC clones 1. Pérez-Vazquez M, Oteo-Iglesias J, Sola-Campoy PJ, contribute to an extended hospital outbreak. Front Microbiol. Carrizo-Manzoni H, Bautista V, Lara N, et al.; Spanish 2019;10:2767. https://doi.org/10.3389/fmicb.2019.02767 Antibiotic Resistance Surveillance Program Collaborating 10. Becker L, Kaase M, Pfeifer Y, Fuchs S, Reuss A, von Laer A, Group. Characterization of carbapenemase-producing et al. Genome-based analysis of carbapenemase-producing Klebsiella oxytoca in Spain, 2016–2017. Antimicrob Agents Klebsiella pneumoniae isolates from German hospital patients, Chemother. 2019;63:e02529–18. https://doi.org/10.1128/ 2008–2014. Antimicrob Resist Infect Control. 2018;7:62. AAC.02529-18 https://doi.org/10.1186/s13756-018-0352-y 2. Arana DM, Ortega A, González-Barberá E, Lara N, 11. Espinal P, Miró E, Segura C, Gómez L, Plasencia V, Coll P, et al. First description of bla carried on an IncX4 plasmid Bautista V, Gómez-Ruíz D, et al.; Spanish Antibiotic NDM-7 Resistance Surveillance Programme Collaborating Group. in Escherichia coli ST679 isolated in Spain. Microb Drug Carbapenem-resistant Citrobacter spp. isolated in Spain from Resist. 2018;24:113–9. https://doi.org/10.1089/mdr.2017.0039 2013 to 2015 produced a variety of carbapenemases 12. Villa J, Carretero O, Viedma E, Lora-Tamayo J, Mingorance J, including VIM-1, OXA-48, KPC-2, NDM-1 and VIM-2. J Chaves F. Emergence of NDM-7-producing multi-drug- Antimicrob Chemother. 2017;72:3283–7. https://doi.org/ resistant Enterobacter hormaechei sequence type ST-78 in 10.1093/jac/dkx325 Spain: a high-risk international clone. Int J Antimicrob 3. Del Barrio-Tofiño E, Zamorano L, Cortes-Lara S, Agents. 2019;53:533–4. https://doi.org/10.1016/ López-Causapé C, Sánchez-Diener I, Cabot G, et al.; j.ijantimicag.2018.11.009 GEMARA-SEIMC/REIPI Pseudomonas study Group. 13. Hamidian M, Nigro SJ. Emergence, molecular mechanisms Spanish nationwide survey on Pseudomonas aeruginosa and global spread of carbapenem-resistant Acinetobacter antimicrobial resistance mechanisms and epidemiology. baumannii. Microb Genom. 2019;5:5. https://doi.org/ J Antimicrob Chemother. 2019;74:1825–35. https://doi.org/ 10.1099/mgen.0.000306 10.1093/jac/dkz147 14. Fernández-Cuenca F, Pérez-Palacios P, Galán-Sánchez F, 4. Nowak J, Zander E, Stefanik D, Higgins PG, Roca I, Vila J, López-Cerero L, López-Hernández I, López Rojas R, et al. First identification of bla carbapenemase in bla - et al.; MagicBullet Working Group WP4. High incidence of NDM-1 OXA-94 pandrug-resistant Acinetobacter baumannii isolates collected producing Acinetobacter baumannii ST85 in Spain. Enferm from patients with ventilator-associated in Infecc Microbiol Clin. 2020;38:11–5. https://doi.org/10.1016/ Greece, Italy and Spain as part of the MagicBullet clinical j.eimc.2019.03.008 trial. J Antimicrob Chemother. 2017;72:3277–82. https://doi.org/10.1093/jac/dkx322 Address for correspondence: I. López-Hernández, Department of 5. García-Castillo M, García-Fernández S, Gómez-Gil R, Microbiology, School of Medicine, Av Sánchez Pizjuán s/n, 41009, Pitart C, Oviaño M, Gracia-Ahufinger I, et al.; iCREST Seville, Spain; email: [email protected] Study Group. Activity of ceftazidime-avibactam against

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Article DOI: https://doi.org/10.3201/eid2609.191772

Carbapenemase-Producing Gram Negative Bacteria in Andalusia, Spain, 2014–2018 Appendix

Appendix Table. Distribution of carbapenemase and STs among the 1243 isolates included in the study Carbapenemase VIM VIM IMP IMP IMP IMP NDM NDM NDM KPC KPC KPC IMI OXA OXA OXA OXA OXA OXA Species Clone -1 -2 -8 -16 -23 -22 -1 -7 -5 -3 -2 -31 -1 -48 -245 -181 -23 -58 -24/40 Total Klebsiella ST525 6 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 6 pneumoniae ST111 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST37 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST307 1 0 0 0 0 0 4 3 0 1 3 0 0 22 0 0 0 0 0 34 ST17 1 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 3 ST22 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST395 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 2 ST348 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST36 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST25 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST514 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST2856 1 0 0 0 0 0 0 0 0 0 0 0 0 3 0 0 0 0 0 4 ST15 11 0 0 0 0 0 0 0 0 0 0 0 0 66 3 0 0 0 0 80 ST128 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST11 1 0 0 0 0 0 0 35 0 0 0 0 0 41 4 0 0 0 0 81 ST392 0 0 1 0 0 0 0 0 0 0 0 0 0 19 0 0 0 0 0 20 ST202 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 ST13 1 0 0 0 0 0 0 0 0 0 0 0 0 5 0 0 0 0 0 6 ST321 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST152 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 1 ST1079 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST1308 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST464 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST512 0 0 0 0 0 0 0 0 0 209 0 1 0 0 0 0 0 0 0 210 ST258 0 0 0 0 0 0 0 0 0 31 0 0 0 0 0 0 0 0 0 31 ST753 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST147 0 0 0 0 0 0 0 0 0 5 1 0 0 8 0 0 0 0 0 14 ST836 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 1 ST899 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST1 0 0 0 0 0 0 0 0 0 0 0 0 0 3 2 0 0 0 0 5

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Carbapenemase VIM VIM IMP IMP IMP IMP NDM NDM NDM KPC KPC KPC IMI OXA OXA OXA OXA OXA OXA Species Clone -1 -2 -8 -16 -23 -22 -1 -7 -5 -3 -2 -31 -1 -48 -245 -181 -23 -58 -24/40 Total ST431 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST251 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 1 ST974 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST405 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 1 ST391 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST225 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST34 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 ST16 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST1963 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 ST981 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST340 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 1 ST1732 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 1 ST2262 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 2 ST3358 0 0 0 0 0 0 0 0 0 0 3 0 0 0 0 0 0 0 0 3 ST3362 0 0 0 0 0 0 0 0 0 0 0 0 0 9 0 0 0 0 0 9 ST1562 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 ST846 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST3360 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 2 ST442 0 0 0 0 0 0 0 0 0 0 0 0 0 5 0 0 0 0 0 5 ST716 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 NA 0 0 0 0 0 0 0 0 0 2 0 0 0 8 0 0 0 0 0 10 K. oxytoca ST145 24 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 25 ST36 14 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 14 ST170 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 ST27 5 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 5 ST213 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST202 22 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 24 ST206 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST82 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST214 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST95 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 NA 9 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 9 K. aerogenes ST93 0 0 0 0 0 0 0 0 0 0 0 0 0 3 0 0 0 0 0 3 ST153 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 Escherichia coli ST1193 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 1 ST145 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST348 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST8327 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST141 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 1 ST88 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST410 0 0 0 0 0 0 0 0 2 0 3 0 0 0 0 0 0 0 0 5 ST617 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 1 ST69 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST1340 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST12 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 NA 5 0 0 0 0 0 0 0 0 0 0 0 0 9 1 0 0 0 0 15 ST78 19 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 21

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Carbapenemase VIM VIM IMP IMP IMP IMP NDM NDM NDM KPC KPC KPC IMI OXA OXA OXA OXA OXA OXA Species Clone -1 -2 -8 -16 -23 -22 -1 -7 -5 -3 -2 -31 -1 -48 -245 -181 -23 -58 -24/40 Total Enterobacter ST111 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 cloacae ST932 0 0 0 0 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 2 ST90 2 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 3 ST114 2 0 1 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 5 ST528 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST96 0 0 0 0 0 1 0 0 0 1 0 0 0 0 0 0 0 0 0 2 ST50 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST413 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST50 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST242 3 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 3 ST24 0 0 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 ST110 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 ST991 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 ST74 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 ST108 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST32 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 1 ST1107 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST113 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST901 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 1 NA 2 0 0 0 0 0 0 0 0 0 1 0 0 2 0 0 0 0 0 5 Citrobacter freundii ST8 1 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 ST22 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST44 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST63 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST170 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST408 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST410 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 ST112 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 2 ST261 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 ST425 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 NA 1 0 0 0 0 0 0 0 0 0 1 0 0 0 1 0 0 0 0 3 C. amalonatycus NA 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 Pseudomonas ST253 0 0 0 9 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 9 aeruginosa ST348 0 0 14 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 14 ST175 0 28 0 0 7 0 0 0 0 0 0 0 0 0 0 0 0 0 0 35 ST308 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 NA 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 P. putida ST15 5 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 5 NA 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 Acinetobacter ST307 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 15 0 0 15 baumannii ST85 0 0 0 0 0 0 3 0 0 0 0 0 0 0 0 0 8 0 0 11 ST2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 170 83 49 302 ST98 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 21 0 21 ST829 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 1 ST745 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 55 0 56 ST32 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 ST164 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 0 0 2

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Carbapenemase VIM VIM IMP IMP IMP IMP NDM NDM NDM KPC KPC KPC IMI OXA OXA OXA OXA OXA OXA Species Clone -1 -2 -8 -16 -23 -22 -1 -7 -5 -3 -2 -31 -1 -48 -245 -181 -23 -58 -24/40 Total NA 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 11 1 4 16 Acinetobacter spp. ST119 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 1 NA 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 Total 167 30 24 9 7 1 12 40 4 249 22 1 1 236 17 1 208 160 54 1243

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Appendix Figure. Geographic distribution of carbapenemase types.

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