Skip to main content
Download PDF

Successful termination of a multi-year wastewater-associated outbreak of NDM-5-carrying E. coli in a hemato-oncological center

Antimicrobial Resistance & Infection Control volume 14, Article number: 27 (2025) Cite this article

Abstract

Background

In May 2018, an outbreak of NDM-5-carrying Escherichia coli (NDM-5-EC) was detected at the hemato-oncology department of a tertiary care center in Austria. This report details the outbreak investigation, control measures and the whole genome sequencing (WGS) data of the outbreak isolates.

Methods

A total of 15 isolates (seven clinical isolates from allogenic stem cell transplant (SCT) recipients and eight wastewater isolates recovered from patients’ toilets) were analyzed by whole genome sequencing.

Results

Genome based typing identified two clusters of the high risk clones ST167/CT12607 and ST617/CT2791. Long-read sequencing of selected isolates from both clusters identified two different plasmids, however with a highly similar genetic context of the blaNDM-5 containing region. Genomic analysis revealed the presence of additional resistance genes, including blaCTX-M-15, and blaOXA-1, and virulence factors. Four patients were colonized with NDM-5-EC, two patients suffered bacteremia caused by the outbreak strain and two deaths were associated with an NDM-5-EC infection. The outbreak source was traced to toilet sewage pipes, which remained persistently contaminated despite extensive cleaning and disinfection. Successful eradication of NDM-5-EC from the installations required disassembly, hot water pressure washing of the sewage pipes and complete replacement of all movable parts. Additionally, colonized patients were instructed to use wheeled commodes instead of toilets, and a pre-admission screening strategy was implemented for all patients undergoing hematologic stem cell transplantation. The outbreak was successfully terminated in November 2020.

Conclusion

NDM-5-EC, especially high-risk clones such as ST167 and ST617, can persist in hospital wastewater systems despite cleaning and disinfection efforts and can cause prolonged outbreaks. Therefore, a comprehensive bundle of interventions like the ones applied in our study is essential, especially in clinical settings with heavily immunosuppressed patients.

Background

The rapid emergence and dissemination of New Delhi metallo-β-lactamase (NDM-5) producing Escherichia coli (NDM-5-EC) poses a significant threat to public health (1). NDM-5, a variant of the NDM enzyme first identified in 2009 (2), confers resistance to a broad spectrum of β-lactam antibiotics, including carbapenems, which are considered last-resort treatments for multidrug-resistant bacterial infections. The NDM-5 carbapenemase has been increasingly reported in Enterobacterales worldwide (3,4,5,6,7) and NDM-5-EC has been implicated in numerous hospital outbreaks (4, 8). These outbreaks frequently involve high-risk clones of sequence types ST167, ST405, ST410, ST617 and ST648, which are known for their capacity to spread rapidly (9, 10). The genetic adaptability of blaNDM-5, often carried on plasmids, facilitates its horizontal transfer between different bacterial species, thereby enhancing its dissemination potential (11).

In this study, we described a wastewater-associated outbreak of NDM-5-EC in a stem cell transplant unit at a tertiary care center in Austria. By integrating comprehensive genomic data with epidemiological investigations, we aimed to enhance our understanding of the mechanisms driving the spread of NDM-5-EC and inform effective strategies for controlling its dissemination in the hospital environment.

Methods

Department description

The affected department at a tertiary care center comprises a general oncology ward (16 rooms for 33 patients), a leukemia and autologous stem cell transplantation (SCT) ward (eight rooms for 12 patients), an allogenic SCT unit (five single rooms) and an outpatient clinic. Its bed occupancy rate ranges between 90 and 100%. The outbreak affected the SCT and the general oncology ward.

Outbreak description

In May 2018, a surveillance stool culture from a hemato-oncological patient (A) and a blood culture from another patient (B) were positive for an NDM-carrying E. coli. A third patient (C) had an NDM-carrying E. coli positive surveillance stool culture in the end of August 2018 followed by a positive blood culture in September 2018. Suspecting a reservoir in the sanitary installations, water samples were taken from sinks, shower drains and toilets in the affected patients’ rooms. NDM-positive E. coli was first found in toilet wastewater in September 2018 and thereafter repeatedly in a total of six patient room toilets on two wards.

The fourth patient (D) had a positive surveillance stool culture in September 2018. A fifth patient (E) had a positive surveillance urine culture in November 2018. In July 2019, a positive surveillance stool culture from a sixth patient (F) was detected, and the seventh and last patient (G) had a positive surveillance urine culture in November 2019.

Case definition

A case was defined as any patient with NDM-EC identified from May 2018 onwards. An ongoing surveillance program (stool, urine, throat swab cultures) had shown no cases before this date. Cases were included in this study until the end of November 2020, when the outbreak was declared terminated.

Patients

Demographic and epidemiological data such as patient outcome (death/survival), stem cell transplant date, microbiology test results and other parameters were collected by chart review, anonymized and then analyzed using Microsoft Excel 2016.

Culture based screening of patients

All neutropenic patients were screened by throat swab, stool culture, urine culture and blood culture twice weekly with a focus on detection of VRE, MRSA and gram-negative bacteria using the following agars: Tryptone Soya Agar with 5% sheep blood, MacConkey Agar, Brilliance ESBL/Brilliance CRE BiPlate, Brilliance VRE (all Thermo Fisher Scientific, UK) and CHROMID MRSA (Biomérieux, France).

Environmental isolates

During the outbreak, environmental isolates were collected by plating 1 ml of toilet trap water on Tryptone Soya Agar with 5% sheep blood and on MacConkey Agar. Additionally, 10 ml toilet trap water was filtrated using the Microfil funnel and filter system (Merck Millipore, UK) with a pore size of 0.45 µm and the filters were placed on plates for culture. Only eight environmental isolates from random time points were archived.

Identification and susceptibility testing

Identification of presumptive Enterobacterales was done by Maldi TOF (Bruker Daltonics, Hilden, Germany) and susceptibility testing according to EUCAST was applied. Isolates meeting screening cut-off values for carbapenemase-producing Enterobacterales (12) and/or growing on the Brilliance ESBL/Brilliance CRE BiPlate were further characterized by PCR.

Bacterial isolates and carbapenemase gene PCR

These isolates were analyzed by conventional PCR with previously published primer sets covering the most prevalent ESBL and carbapenemase genes (13, 14).

The first clinical isolates from all seven patients (labelled A-G) and eight isolates (labelled 1–1 to 6–1) obtained from toilet wastewater of six different patient rooms (one toilet was sampled three times over one year) were included in the study. Isolates (n = 15) were retrieved from cryobanks (Mast, Reinfeld, Germany) and cultured on Tryptone Soya Agar with 5% sheep blood for whole genome sequencing (WGS).

DNA isolation, WGS and typing

DNA from overnight cultures was extracted using the DNeasy UltraClean Microbial Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions. DNA libraries were prepared using the Illumina DNA Prep M Tagmentation Kit combined with IDT for Illumina UD Indexes (Illumina, San Diego, USA) followed by whole-genome sequencing on the iSeq 100 system (Illumina, San Diego, USA) with 2 × 150 bp paired end reads. Bioinformatic analyses were performed with Ridom SeqSphere + software v9.0.8 (Ridom GmbH, Münster, Germany) (15). FastQC v0.11.9 was used for quality analysis of the sequences (16). Raw reads were assembled with SKESA v2.4.0 (17) and assembly remapping and polishing was performed with the BWA-MEM v0.7.15 algorithm (18).

NCBI AMRFinderPlus 3.11.2 was used for identification of resistance genes (19). Likewise, Warwick MLST (20) and cgMLST (Enterobase) (21) analyses were run on SeqSphere + and a minimum spanning tree (MST) was generated using the default threshold of ≤ 10 allelic differences (AD). Genomes were additionally analysed using VirulenceFinder-2.0 Server with default settings (Software version: 2.0.5 (2024–01-31), Database version: (2022–12-02)) (22).

For further investigation of the location of blaNDM-5 in the bacterial genome, four selected isolates (patient C, patient F and two environmental isolates: room 2–1 and room 6–1) additionally underwent long-read sequencing with Oxford Nanopore Technologies. Genome libraries were prepared using the rapid barcoding kit SQK-RBK004 (ONT, Oxford, United Kingdom) according to the standard protocol and loaded on a R9.4.1 flowcell (FLO-MIN106D, ONT, Oxford, United Kingdom). Sequencing was performed on a MinION M1kB for 72 h. Quality check and fast basecalling were done with MinKNOW and Guppy v6.3.5 software. Hybrid assembly was performed using unicycler v. 0.5 with default parameters. For plasmid visualization and comparison DNA Features Viewer v. 3.1.2 was used (23). Plasmids were annotated for resistance genes using NCBI-AMRFinder version 3.12.8 (db version 2024–05-02.2) (24).

Results

Patients

A total of seven allo-SCT patients were affected by this outbreak, with their demographic and epidemiological characteristics summarized in Table 1.

Table 1 Characteristics of the seven allogenic stem cell transplant recipients affected by the outbreak (GvHD: graft versus host disease, SCT: stem cell transplant)
Full size table

Figure 1 shows the epidemiological curve of the outbreak by collection date of the first NDM-5-EC isolate from each patient. Most patients had multiple positive samples, typically collected from stool, urine or throat swabs as part of routine screening, four of them were considered colonized. Patient B and C had NDM-5-EC bacteremia and patient B and E did not survive the infection. Three additional patients died of unrelated causes during follow-up. Two patients are still alive and persistently colonized, as shown by screening of stool samples during follow-up until the time of this publication.

Fig. 1
figure 1

Epidemiological curve representing the seven patients (A-G) with NDM-5-EC and their occupation of the affected rooms within the outbreak period from May 2018 to December 2019. Only first detections are shown. Additionally, sampling dates for the environmental isolates (denoted in blue) and infection control events are shown (*: disinfection of toilet bowl with bleach and UV light, increased frequency of cleaning and disinfection of patient-near surfaces, #: change of toilet rubber cuff, °: weekly application of bleach in toilet trap water until negative cultures, §: hot water pressure washing and change of movable parts)

Full size image

Isolate susceptibility data

All isolates were multidrug resistant according to the definition of the ECDC (25). Detailed results of antimicrobial susceptibility testing are presented in Table 2.

Table 2 Antimicrobial susceptibility testing data according to EUCAST for all patient and environmental isolates. Minimum inhibitory concentrations are indicated in brackets. n.d.: not determined
Full size table

Outbreak control measures

When the movement of patients in the affected wards was tracked, it was found that patients A and B had consecutively occupied the same room (Room 1). A screening regimen for neutropenic patients (twice weekly rectal swab/stool sample, urine, throat swab) was already in place and was continued with special regard to NDM carrying E. coli. Additionally, a pre-admission screening strategy was implemented for all patients undergoing hematologic stem cell transplantation. Single-room care for colonized patients with strict contact precautions and increased frequency of cleaning and disinfection of patient-near surfaces especially in bathrooms was initiated after detection of the first case.

In September 2018, after detection of patient C and D (who had also occupied Room 1), a source in the patient room was suspected. Following reports describing hospital water as a reservoir for carbapenemase producing organisms causing infections (26), analysis of water samples revealed contamination of the pipes draining the toilets. Consequently, a vigorous cleaning routine was established: All affected toilets were intensively cleaned, and weekly application of 500 ml of household bleach (2.8 g sodium hypochlorite per 100 g solution) was initiated. Weekly, and later monthly, surveillance of wastewater from the affected toilets was implemented. Toilets were cleared for use only after three negative surveillance cultures. However, due to the repeated reappearance of the outbreak strain, especially in Room 1, and ongoing transmission to patients, four of the affected toilets could only be released for use after the total renewal of all movable parts (toilet bowl, rubber seal, closet flange) and intense cleaning of the drainpipes with hot water pressure washing (Fig. 2).

Fig. 2
figure 2

Hot water high pressure cleaning of the drainpipes after complete removal of the toilet and view of the drainpipe biofilm before cleaning

Full size image

Ultimately, colonized patients were discouraged from using the toilets and were instead advised to use wheeled commode chairs without connection to the hospital wastewater system for the total length of their hospital stay to prevent re-contamination of the installations.

The toilet installations were finally cleared from NDM-5-EC in June 2020.

WGS-based typing

Two different sequence types / cgMLST complex types were assigned to the outbreak isolates (ST617/CT12607 n = 12, ST167/CT2791 n = 3). A cgMLST analysis grouped the 15 isolates into two clusters (threshold: ≤ 10 AD) corresponding to the two STs, respectively (Fig. 3). The two clusters differed by 700 alleles and isolates within the clusters differed by a maximum of two and three alleles, respectively (Fig. 3). Table 3 provides an overview of assigned STs and cgMLST CTs for all isolates. They shared a common betalactam resistance gene pattern (blaNDM-5, blaCTX-M-15, blaOXA-1) with variable detection of blaTEM-1 and multiple resistance genes for other antibiotic classes (Table 3). Moreover, a wide array of virulence genes as well as genes associated with disinfectant tolerance and biofilm formation was detected (Table 3).

Fig. 3
figure 3

Minimum spanning tree showing the two clusters detected in the outbreak. ST167 isolates are colored blue, ST617 isolates are colored red

Full size image
Table 3 Whole genome sequencing data for all isolates. Data is sorted by sampling date, isolates showing the cgMLST 2791 complex type are colored light grey.
Full size table

Analysis of the hybrid assemblies from isolates of both clusters (patient C (ST167/ CT2791), patient F (ST617/ CT12607), room 2–1 (ST617/ CT12607) and room 6–1 (ST167/ CT2791)) revealed the localization of blaNDM-5 on IncFII type plasmids (Fig. 4, image generated with DNA Features Viewer). The plasmids were of different sizes: Plasmid 1 measured approximately 133–139 kb (patient C and room 6–1) and Plasmid 2 approximately 168 kb (patient F and room 2–1). Plasmids of similar size were highly identical over large collinear blocks (Fig. 4), while Plasmid 1 and Plasmid 2 were approximately 97% identical (74% coverage). All plasmids shared common identical blocks encoding blaNDM-5 on a 13,801 bp long identical segment. The recombined regions around blaNDM-5 were interspersed with multiple copies of the insertion sequence IS26.

Fig. 4
figure 4

Comparison of common identical nucleotide blocks between the plasmids of four isolates. The image was created using DNA Features Viewer. A minimum length of 1000 bp was required for a common block. Location of resistance conveying genes was added. Patient C and Room 6–1 (NDM-5-EC ST167) plasmids share large common blocks (block 2 and 3), while Patient F and room 2–1 (NDM-5-EC ST617) plasmids share a common plasmid backbone (block 1 and 4). Some resistance gene cassettes, including blaNDM-5, are common to all four plasmids (block 5 and 6)

Full size image

Discussion

In this study, we report a wastewater-associated NDM-5-EC outbreak in an Austrian hospital, which led to colonization as well as infection of allogenic stem cell (allo-SCT) transplant recipients. This patient population is particularly vulnerable due to bone marrow suppression, impaired mucosal barriers and often prolonged hospitalization with exposure to broad-spectrum antibiotics (such as antipseudomonal cephalosporins and carbapenems) during therapy of neutropenic fever (27). Colonization with multidrug resistant bacteria such as NDM-5-EC has been shown to pose a risk for subsequent invasive infection in allo-SCT recipients and negatively impact survival (28). Therefore, prompt detection of colonized patients through screening is recommended to minimize onward transmission (27).

Hospitals are critical reservoirs for multidrug-resistant bacteria due to the high antimicrobial selection pressure, especially in wards treating heavily immunocompromised patients (29). Antibiotics excreted by patients are discharged into the hospital wastewater system, where they may reach high concentrations (30), exerting selection pressure on environmental bacteria and promoting the emergence and spread of multidrug-resistant clones (31). Several studies have investigated the role of toilets and sanitary installations in the transmission of carbapenemase-producing bacteria within hospital settings. A recent paper covered a prolonged outbreak of carbapenemase-producing K. pneumoniae and P. aeruginosa, finding that environmental and clinical isolates were closely related (29). An outbreak of NDM-producing ST167 E. coli was linked to a toilet in a Danish hospital where environmental samples from the toilet water trap revealed the presence of the outbreak isolate, indicating a possible source of infection (4). Similarly, in the burn center of Ghent University Hospital, an outbreak of OXA-48-producing Klebsiella pneumoniae was traced to toilet and drain water. The outbreak strain persisted in some rooms even after two months of daily disinfection with bleach, highlighting the potential of the strain to spread between rooms through common wastewater plumbing and the failure of disinfectants to prevent recolonization after discontinuation (32). This corresponds to our experience of continuing reappearance of NMD-5-EC in affected toilets until complete replacement of the affected installations, underscoring the importance of targeted infection control measures and environmental disinfection to mitigate the risk of pathogen transmission through hospital sanitary facilities, especially if they encode diverse persistence and resistance mechanisms.

Additionally, the characteristics of the outbreak strains possibly contributed to the extended duration of this outbreak. The initially obtained short-read sequencing data revealed two separate clusters, which could have been interpreted as two distinct outbreak events. However, due to the very low prevalence of NDM-5-EC in Austria (33), we decided to perform further analysis by Nanopore sequencing. As for the STs identified here, both ST167 as well as ST617 E. coli emerged from the ST10 clonal complex and are known for causing outbreaks due to unique virulence and surface antigen features (34), which were also detected in our isolates. They are also carriers of NDM-type carbapenemases and are often isolated in healthcare settings (35,36,37). In our case, transmission of blaNDM-5 between the two different E. coli sequence types via plasmid exchange may be assumed, as the long-read sequencing data demonstrate a highly similar genetic context of blaNDM-5 in all four analyzed isolates despite the differences between the detected plasmids. The NDM gene is commonly located within recombination-prone and transposon-rich genomic regions (38) which is consistent with our findings. Indeed, recombination between multiple plasmid types carrying blaNDM-5 has been well documented in various bacterial species, even in a single patient (39, 40). Additionally, plasmid DNA has been shown to remain viable for several days in aqueous environments with favorable conditions for biofilm formation (41), and long-distance dissemination of antimicrobial resistance genes via outer membrane vesicles has recently been described (42, 43). These factors suggest that horizontal transfer of the NMD-5 gene between the two E. coli STs could have occurred within the hospital’s wastewater systems. Nevertheless, the overall differences observed between the plasmids in the two sequence types raise the possibility of a second independent introduction of NDM-5-EC during this outbreak, although this remains epidemiologically unlikely regarding the rarity of such strains in Austria (44).

Taken together, all these factors contributed to the difficulties in terminating this outbreak. Only a combination of measures covering installations (complete exchange of parts, high-pressure cleaning, chlorination), infection control (single room patient care with PPE, screening) and patient behavior (education, use of commodes for colonized patients) was successful.

Conclusions

Detection of wastewater-associated NDM-5-EC in a low-incidence setting should prompt aggressive outbreak management, especially in high-risk wards such as hemato-oncology. The study demonstrated that traditional disinfection protocols were insufficient to eliminate persistent contamination in wastewater pipes, necessitating mechanical interventions such as disassembly and high-pressure hot water cleaning, followed by infrastructure replacements. Further research including long-term genomic surveillance is essential to explore the mechanisms of plasmid exchange and resistance gene dissemination in hospital environments. Additionally, optimized products and protocols to prevent contamination of sanitary installations are needed to prevent the spread of antibiotic-resistant pathogens in healthcare facilities.

Availability of data and materials

This Whole Genome Shotgun project has been deposited at the Sequence Read Archive (SRA) under the accession no. PRJNA1165667 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1165667). The version described in this paper is the first version.

Abbreviations

AD:

Allelic Differences

bp:

Base Pairs

cgMLST:

Core genome Multilocus Sequence Typing

CT:

Complex type

DNA:

Deoxyribonucleic Acid

E. coli (EC):

Escherichia coli

GvHD:

Graft versus host disease

MLST:

Multilocus Sequence Typing

MRSA:

Methicillin resistant Staphylococcus aureus

MST:

Minimum Spanning Tree

NDM:

New Delhi Metallo-beta-lactamase

ONT:

Oxford Nanopore Technologies

PCR:

Polymerase chain reaction

SCT:

Stem Cell Transplant

ST:

Sequence type

VRE:

Vancomycin resistant Enterococci

WGS:

Whole Genome Sequencing

References

  1. Linkevicius M, Bonnin RA, Alm E, Svartström O, Apfalter P, Hartl R, et al. Rapid cross-border emergence of NDM-5-producing escherichia coli in the European Union/European Economic Area, 2012 to June 2022. Eurosurveillance. 2022. https://doiorg.publicaciones.saludcastillayleon.es/10.2807/1560-7917.ES.2023.28.19.2300209.

    Article  PubMed  PubMed Central  Google Scholar 

  2. Yong D, Toleman MA, Giske CG, Cho HS, Sundman K, Lee K, et al. 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. 2009;53(12):5046–54.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Hammerum AM, Hansen F, Olesen B, Struve C, Holzknecht BJ, Andersen PS, et al. Investigation of a possible outbreak of NDM-5-producing ST16 Klebsiella pneumoniae among patients in Denmark with no history of recent travel using whole-genome sequencing. J Global Antimicrob Res. 2015;3(3):219–21.

    Article  Google Scholar 

  4. Andrews V, Hasman H, Midttun M, Feldthaus MB, Porsbo LJ, Holzknecht B et al. A hospital outbreak of an NDM-producing ST167 E. coli with a possible link to a toilet. Journal of Hospital Infection 2021.

  5. Hans JB, Pfennigwerth N, Neumann B, Pfeifer Y, Fischer MA, Eisfeld J, et al. Molecular surveillance reveals the emergence and dissemination of NDM-5-producing Escherichia coli high-risk clones in Germany, 2013 to 2019. Euro Surveill. 2023. https://doiorg.publicaciones.saludcastillayleon.es/10.2807/1560-7917.ES.2023.28.10.2200509.

    Article  PubMed  PubMed Central  Google Scholar 

  6. Adler A, Ghosh H, Gross A, Rechavi A, Lasnoy M, Assous MV, et al. Molecular features and transmission of NDM-producing Enterobacterales in Israeli hospitals. J Antimicrob Chemother. 2023;78(3):719–23.

    Article  CAS  PubMed  Google Scholar 

  7. Chakraborty T, Sadek M, Yao Y, Imirzalioglu C, Stephan R, Poirel L, et al. Cross-Border Emergence of Escherichia coli Producing the Carbapenemase NDM-5 in Switzerland and Germany. J Clin Microbiol. 2021. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/JCM.02238-20.

    Article  PubMed  PubMed Central  Google Scholar 

  8. Bibbolino G, Di Lella FM, Oliva A, Lichtner M, Del Borgo C, Raponi G, et al. Molecular epidemiology of NDM-5-producing Escherichia coli high-risk clones identified in two Italian hospitals in 2017–2019. Diagn Microbiol Infect Dis. 2021;100(4): 115399.

    Article  CAS  PubMed  Google Scholar 

  9. Barrado L, Pérez-Vázquez M, Del Pozo JL, Martín-Salas C, Leiva J, Mazón A, et al. Clonal transmission of NDM-5-producing escherichia coli belonging to high-risk sequence type ST405. Int J Antimicrob Agents. 2018;52(1):123–4.

    Article  CAS  PubMed  Google Scholar 

  10. Bitar I, Piazza A, Gaiarsa S, Villa L, Pedroni P, Oliva E, et al. ST405 NDM-5 producing escherichia coli in Northern Italy: the first two clinical cases. Clin Microbiol Infect. 2017;23(7):489–90.

    Article  CAS  PubMed  Google Scholar 

  11. Yang QE, Ma X, Zeng L, Wang Q, Li M, Teng L, et al. Interphylum dissemination of NDM-5-positive plasmids in hospital wastewater from Fuzhou, China: a single-centre, culture-independent, plasmid transmission study. Lancet Microbe. 2024;5(1):e13-23.

    Article  PubMed  Google Scholar 

  12. EUCAST subcommittee for detection of resistance mechanisms. EUCAST guideline for the detection of resistance mechanisms and specific resistances of clinical and/or epidemiological importance; 2017 [cited 2025 Feb 25]. Available from: URL: https://www.eucast.org/resistance_mechanisms.

  13. Poirel L, Walsh TR, Cuvillier V, Nordmann P. Multiplex PCR for detection of acquired carbapenemase genes. Diagn Microbiol Infect Dis. 2011;70(1):119–23.

    Article  CAS  PubMed  Google Scholar 

  14. Dallenne C, Da Costa A, Decré D, Favier C, Arlet G. Development of a set of multiplex PCR assays for the detection of genes encoding important beta-lactamases in Enterobacteriaceae. J Antimicrob Chemother. 2010;65(3):490–5.

    Article  CAS  PubMed  Google Scholar 

  15. Jünemann S, Sedlazeck FJ, Prior K, Albersmeier A, John U, Kalinowski J, et al. Updating benchtop sequencing performance comparison. Nat Biotechnol. 2013;31(4):294–6.

    Article  PubMed  Google Scholar 

  16. https://www.bioinformatics.babraham.ac.uk/projects/fastqc; 2021 [cited 2025 Feb 24]. Available from: URL: https://www.bioinformatics.babraham.ac.uk/projects/fastqc.

  17. Souvorov A, Agarwala R, Lipman DJ. SKESA: strategic k-mer extension for scrupulous assemblies. Genome Biol. 2018;19(1):153.

    Article  PubMed  PubMed Central  Google Scholar 

  18. Li H. Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM; 2013 Mar 16. Available from: URL: http://arxiv.org/pdf/1303.3997.

  19. Feldgarden M, Brover V, Haft DH, Prasad AB, Slotta DJ, Tolstoy I, et al. Validating the AMRFinder tool and resistance gene database by using antimicrobial resistance genotype-phenotype correlations in a collection of isolates. Antimicrob Agents Chemother. 2019. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/AAC.00483-19.

    Article  PubMed  PubMed Central  Google Scholar 

  20. Wirth T, Falush D, Lan R, Colles F, Mensa P, Wieler LH, et al. Sex and virulence in Escherichia coli: an evolutionary perspective. Mol Microbiol. 2006;60(5):1136–51.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Zhou Z, Alikhan N-F, Mohamed K, Fan Y, Achtman M. The EnteroBase user’s guide, with case studies on Salmonella transmissions, Yersinia pestis phylogeny, and Escherichia core genomic diversity. Genome Res. 2020;30(1):138–52.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Joensen KG, Scheutz F, Lund O, Hasman H, Kaas RS, Nielsen EM, et al. Real-time whole-genome sequencing for routine typing, surveillance, and outbreak detection of verotoxigenic Escherichia coli. J Clin Microbiol. 2014;52(5):1501–10.

    Article  PubMed  PubMed Central  Google Scholar 

  23. Zulkower V, Rosser S. DNA Features Viewer: a sequence annotation formatting and plotting library for Python. Bioinformatics. 2020;36(15):4350–2.

    Article  CAS  PubMed  Google Scholar 

  24. Feldgarden M, Brover V, Gonzalez-Escalona N, Frye JG, Haendiges J, Haft DH, et al. AMRFinderPlus and the Reference Gene Catalog facilitate examination of the genomic links among antimicrobial resistance, stress response, and virulence. Sci Rep. 2021. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41598-021-91456-0.

    Article  PubMed  PubMed Central  Google Scholar 

  25. Magiorakos A-P, Srinivasan A, Carey RB, Carmeli Y, Falagas ME, Giske CG, et al. Multidrug-resistant, extensively drug-resistant and pandrug-resistant bacteria: an international expert proposal for interim standard definitions for acquired resistance. Clin Microbiol Infect. 2012;18(3):268–81.

    Article  CAS  PubMed  Google Scholar 

  26. Kizny Gordon AE, Mathers AJ, Cheong EYL, Gottlieb T, Kotay S, Walker AS, et al. The hospital water environment as a reservoir for carbapenem-resistant organisms causing hospital-acquired infections-a systematic review of the literature. Clin Infect Dis. 2017;64(10):1435–44.

    Article  PubMed  Google Scholar 

  27. Sureda A. The EBMT Handbook: Hematopoietic Cell Transplantation and Cellular Therapies. 8th ed. Cham: Springer International Publishing AG; 2024. Available from: URL: https://ebookcentral.proquest.com/lib/kxp/detail.action?docID=31281792.

  28. Sadowska-Klasa A, Piekarska A, Prejzner W, Bieniaszewska M, Hellmann A. Colonization with multidrug-resistant bacteria increases the risk of complications and a fatal outcome after allogeneic hematopoietic cell transplantation. Ann Hematol. 2018;97(3):509–17.

    Article  CAS  PubMed  Google Scholar 

  29. Neidhöfer C, Sib E, Neuenhoff M, Schwengers O, Dummin T, Buechler C, et al. Hospital sanitary facilities on wards with high antibiotic exposure play an important role in maintaining a reservoir of resistant pathogens, even over many years. Antimicrob Resist Infect Control. 2023;12(1):33.

    Article  PubMed  PubMed Central  Google Scholar 

  30. Voigt AM, Faerber HA, Wilbring G, Skutlarek D, Felder C, Mahn R, et al. The occurrence of antimicrobial substances in toilet, sink and shower drainpipes of clinical units: A neglected source of antibiotic residues. Int J Hyg Environ Health. 2019;222(3):455–67.

    Article  CAS  PubMed  Google Scholar 

  31. Lépesová K, Olejníková P, Mackuľak T, Cverenkárová K, Krahulcová M, Bírošová L. Hospital Wastewater-Important Source of Multidrug Resistant Coliform Bacteria with ESBL-Production. Int J Environ Res Public Health. 2020;17(21):7827.

    Article  PubMed  PubMed Central  Google Scholar 

  32. Heireman L, Hamerlinck H, Vandendriessche S, Boelens J, Coorevits L, de Brabandere E, et al. Toilet drain water as a potential source of hospital room-to-room transmission of carbapenemase-producing Klebsiella pneumoniae. J Hosp Infect. 2020. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.jhin.2020.07.017.

    Article  PubMed  Google Scholar 

  33. Resistenzbericht Österreich AURES 2019: Antibiotikaresistenz und Verbrauch antimikrobieller Substanzen in Österreich. Wien; 2021 [cited 2025 Feb 24]. Available from: URL: https://broschuerenservice.sozialministerium.at/Home/Download?publicationId=290&attachmentName=Resistenzbericht_%C3%96sterreich_AURES_2019_pdfUA.pdf.

  34. Garcia-Fernandez A, Villa L, Bibbolino G, Bressan A, Trancassini M, Pietropaolo V, et al. Novel insights and features of the NDM5Producing escherichia coli sequence type 167 highrisk clone. mSphere. 2020. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/mSphere.00269-20.

    Article  PubMed  PubMed Central  Google Scholar 

  35. Zou H, Han J, Zhao L, Wang Di, Guan Y, Wu T, et al. The shared NDM-positive strains in the hospital and connecting aquatic environments. Sci Total Environ. 2023;860:160404.

    Article  CAS  PubMed  Google Scholar 

  36. Wu W, Feng Y, Tang G, Qiao F, McNally A, Zong Z. NDM Metallo-β-lactamases and their bacterial producers in health care settings. Clin Microbio Rev. 2019. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/CMR.00115-18.

    Article  Google Scholar 

  37. Acman M, Wang R, van Dorp L, Shaw LP, Wang Q, Luhmann N, et al. Role of mobile genetic elements in the global dissemination of the carbapenem resistance gene blaNDM. Nat Commun. 2022;13(1):1131.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Li X, He J, Yu Y, Zhou H, Tu Y, Hua X. Dynamic evolution and inter-species transfer of blaNDM-5 plasmid in vivo in a single patient. Clin Microbiol Infect. 2023;29(2):265–8.

    Article  PubMed  Google Scholar 

  39. Ma Z, Wang B, Zeng D, Ding H, Zeng Z. Rapid dissemination of blandm-5 gene among carbapenem-resistant escherichia coli isolates in a yellow-feather broiler farm via multiple plasmid replicon. Pathogens. 2024;13(5):387.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Winder EM, Bonheyo GT. DNA Persistence in a Sink Drain Environment. PLoS ONE. 2015;10(7): e0134798.

    Article  PubMed  PubMed Central  Google Scholar 

  41. Tang B, Yang A, Liu P, Wang Z, Jian Z, Chen X, et al. Outer Membrane Vesicles Transmitting blaNDM-1 Mediate the Emergence of Carbapenem-Resistant Hypervirulent Klebsiella pneumoniae. Antimicrob Agents Chemother. 2023;67(5): e0144422.

    Article  PubMed  Google Scholar 

  42. Carrera Páez LC, Olivier M, Gambino AS, Poklepovich T, Aguilar AP, Quiroga MP, et al. Sporadic clone Escherichia coli ST615 as a vector and reservoir for dissemination of crucial antimicrobial resistance genes. Front Cell Infect Microbiol. 2024;14:1368622.

    Article  PubMed  PubMed Central  Google Scholar 

  43. European Centre for Disease Prevention and Control. Antimicrobial resistance in the EU/EEA (EARS-Net) - Annual epidemiological report for 2022. Stockholm: ECDC; 2023 [cited 2024 Sep 30]. Available from: URL: https://www.ecdc.europa.eu/en/publications-data/surveillance-antimicrobial-resistance-europe-2022.

Download references

Acknowledgements

The authors thank all members of the infection control team (Marie-Luise Einfalt, Christian Hartl, Petra Roitner, Vera Bogicevic) for their excellent work.

Funding

No external funding was received for this study.

Author information

Authors and Affiliations

  1. National Reference Center for Antimicrobial Resistance, Institute for Hygiene, Microbiology and Tropical Medicine, Ordensklinikum Linz Elisabethinen, Fadingerstraße 1, 4020, Linz, Austria

    Heidrun Kerschner, Linda Jernej, Lucia Berning, Anna Blaimschein, Petra Apfalter & Rainer Hartl

  2. Laboratory of Photodynamic Inactivation of Microorganisms, Department of Biosciences and Medical Biology, Paris Lodron University Salzburg, Hellbrunnerstraße 34, 5020, Salzburg, Austria

    Linda Jernej

  3. Institute of Medical Microbiology and Hygiene, Austrian Agency for Health and Food Safety (AGES), Währingerstraße 25a, 1090, Vienna, Austria

    Adriana Cabal, Patrick Hyden & Werner Ruppitsch

  4. Department of Internal Medicine 1, Ordensklinikum Linz Elisabethinen, Fadingerstraße 1, 4020, Linz, Austria

    Sigrid Machherndl-Spandl

  5. Medical Faculty, Johannes Kepler University Linz, Altenberger Strasse 69, 4040, Linz, Austria

    Heidrun Kerschner & Rainer Hartl

  6. FoodHub – Centre of Excellence for Digitalization of Microbial Food Safety Risk Assessment and Quality Parameters for Accurate Food Authenticity Certification, University of Donja Gorica, Podgorica, Montenegro

    Werner Ruppitsch

Authors
  1. Heidrun Kerschner
    View author publications

    You can also search for this author inPubMed Google Scholar

  2. Linda Jernej
    View author publications

    You can also search for this author inPubMed Google Scholar

  3. Adriana Cabal
    View author publications

    You can also search for this author inPubMed Google Scholar

  4. Patrick Hyden
    View author publications

    You can also search for this author inPubMed Google Scholar

  5. Sigrid Machherndl-Spandl
    View author publications

    You can also search for this author inPubMed Google Scholar

  6. Lucia Berning
    View author publications

    You can also search for this author inPubMed Google Scholar

  7. Anna Blaimschein
    View author publications

    You can also search for this author inPubMed Google Scholar

  8. Werner Ruppitsch
    View author publications

    You can also search for this author inPubMed Google Scholar

  9. Petra Apfalter
    View author publications

    You can also search for this author inPubMed Google Scholar

  10. Rainer Hartl
    View author publications

    You can also search for this author inPubMed Google Scholar

Contributions

Study design: HK, RH, PA Epidemiological study: HK, LJ, SMS Microbiological analysis: LB, AB WGS: LJ, LB, AB, AC, WR, PH Bioinformatics: AC, PH, LB, AB, LJ Data analysis: HK, LJ, AC, WR, PH, SMS Manuscript writing: HK, RH Manuscript revision: PA, WR All authors read and approved the final manuscript. Manuscript revision: PA, WR. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Rainer Hartl.

Ethics declarations

Ethics approval and consent to participate

The study was approved by the ethics committee of the medical faculty of Johannes Kepler University Linz (vote EK1077/2021). Consent to participate was not obtained since the study was done retrospectively and without additional interventions for patients.

Competing interests

The authors declare no competing interests.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material. You do not have permission under this licence to share adapted material derived from this article or parts of it. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc-nd/4.0/.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Kerschner, H., Jernej, L., Cabal, A. et al. Successful termination of a multi-year wastewater-associated outbreak of NDM-5-carrying E. coli in a hemato-oncological center. Antimicrob Resist Infect Control 14, 27 (2025). https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s13756-025-01539-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s13756-025-01539-0

Keywords

Antimicrobial Resistance & Infection Control

ISSN: 2047-2994

Contact us