1887

Microbial Genomics logo

Volume 9, Issue 9

IS-related large-scale deletion of chromosomal regions harbouring the oxygen-insensitive nitroreductase gene causes nitrofurantoin heteroresistance in Open Access

Abstract

Nitrofurantoin is a broad-spectrum first-line antimicrobial used for managing uncomplicated urinary tract infection (UTI). Loss-of-function mutations in chromosomal genes and of are known to reduce nitrofurantoin susceptibility. Here, we report the discovery of nitrofurantoin heteroresistance in clinical isolates and a novel genetic mechanism associated with this phenomenon. Subpopulations with lower nitrofurantoin susceptibility than major populations (hereafter, nitrofurantoin-resistant subpopulations) in two blood isolates (previously whole-genome sequenced) were identified using population analysis profiling. Each isolate was known to have a loss-of-function mutation in . From each isolate, four nitrofurantoin-resistant isolates were derived at a nitrofurantoin concentration of 32 mg l, and a comparator isolate was obtained without any nitrofurantoin exposure. Genomes of derived isolates were sequenced on Illumina and Nanopore MinION systems. Genetic variation between isolates was determined based on genome assemblies and read mapping. Nitrofurantoin minimum inhibitory concentrations (MICs) of both blood isolates were 64 mg l, with MICs of major nitrofurantoin-susceptible populations varying from 4 to 8 mg l. Two to 99 c.f.u. per million demonstrated growth at the nitrofurantoin concentration of 32 mg l, which is distinct from that of a homogeneously susceptible or resistant isolate. Derived nitrofurantoin-resistant isolates had 11–66 kb deletions in chromosomal regions harbouring , and all deletions were immediately adjacent to IS-family insertion sequences. Our findings demonstrate that the IS-associated large-scale genetic deletion is a hitherto unrecognized mechanism of nitrofurantoin heteroresistance and could compromise UTI management. Further, frequencies of resistant subpopulations from nitrofurantoin-heteroresistant isolates may challenge conventional nitrofurantoin susceptibility testing in clinical settings.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): comparative genomics , Escherichia coli , hybrid genome assembly , insertion sequences , IS1 elements , nitrofurantoin heteroresistance , population analysis profiling , resistance mechanism , structural variation and whole-genome sequencing
Funding
This study was supported by the:
  • Rosetrees Trust (Award M683)
    • Principle Award Recipient: ElitaJauneikaite
  • Wellcome Trust (Award PSN109)
    • Principle Award Recipient: YuWan
  • NIHR Imperial Biomedical Research Centre
    • Principle Award Recipient: ShiraneeSriskandan
  • National Institute for Health Research Health Protection Research Unit (Award NIHR200876)
    • Principle Award Recipient: ApplicableNot
  • UK Health Security Agency
    • Principle Award Recipient: YuWan
  • National Institute for Health Research Health Protection Research Unit (Award NIHR200876)
    • Principle Award Recipient: WanYuJauneikaiteElitaBrownColin S.EllingtonMatthew J.SriskandanShiranee
© 2023 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
Loading

Article metrics loading...

/content/journal/mgen/10.1099/mgen.0.001102
2023-09-06
2024-05-03
Loading full text...

Full text loading...

/deliver/fulltext/mgen/9/9/mgen001102.html?itemId=/content/journal/mgen/10.1099/mgen.0.001102&mimeType=html&fmt=ahah

References

  1. National Institute for Health and Care Excellence Guidance, NICE advice and quality standards (NG109, NG112, NG113). NICE 2018 https://www.nice.org.uk/guidance/published accessed 13 March 2023
     [Google Scholar]
  2. Mottaghizadeh F, Mohajjel Shoja H, Haeili M, Darban-Sarokhalil D. Molecular epidemiology and nitrofurantoin resistance determinants of nitrofurantoin-non-susceptible Escherichia coli isolated from urinary tract infections. J Glob Antimicrob Resist 2020; 21:335–339 [View Article] [PubMed]
     [Google Scholar]
  3. Ho P-L, Ng K-Y, Lo W-U, Law PY, Lai EL-Y et al. Plasmid-mediated OqxAB is an important mechanism for nitrofurantoin resistance in Escherichia coli. Antimicrob Agents Chemother 2015; 60:537–543 [View Article] [PubMed]
     [Google Scholar]
  4. Abernethy J, Guy R, Sheridan EA, Hopkins S, Kiernan M et al. Epidemiology of Escherichia coli bacteraemia in England: results of an enhanced sentinel surveillance programme. J Hosp Infect 2017; 95:365–375 [View Article] [PubMed]
     [Google Scholar]
  5. Ny S, Edquist P, Dumpis U, Gröndahl-Yli-Hannuksela K, Hermes J et al. Antimicrobial resistance of Escherichia coli isolates from outpatient urinary tract infections in women in six European countries including Russia. J Glob Antimicrob Resist 2019; 17:25–34 [View Article] [PubMed]
     [Google Scholar]
  6. UK Health Security Agency English surveillance programme for antimicrobial utilisation and resistance (ESPAUR) Report 2021 to 2022 Annexe 2022 https://assets.publishing.service.gov.uk/government/uploads/system/uploads/attachment_data/file/1118730/ESPAUR-report-2021-2022-annexe.pdf accessed 13 March 2023
     [Google Scholar]
  7. Andersson DI, Nicoloff H, Hjort K. Mechanisms and clinical relevance of bacterial heteroresistance. Nat Rev Microbiol 2019; 17:479–496 [View Article] [PubMed]
     [Google Scholar]
  8. El-Halfawy OM, Valvano MA. Antimicrobial heteroresistance: an emerging field in need of clarity. Clin Microbiol Rev 2015; 28:191–207 [View Article] [PubMed]
     [Google Scholar]
  9. Band VI, Crispell EK, Napier BA, Herrera CM, Tharp GK et al. Antibiotic failure mediated by a resistant subpopulation in Enterobacter cloacae. Nat Microbiol 2016; 1:16053 [View Article] [PubMed]
     [Google Scholar]
  10. Wan Y, Mills E, Leung RCY, Vieira A, Zhi X et al. Alterations in chromosomal genes nfsA, nfsB, and ribE are associated with nitrofurantoin resistance in Escherichia coli from the United Kingdom. Microb Genom 2021; 7:000702 [View Article]
     [Google Scholar]
  11. Jauneikaite E, Honeyford K, Blandy O, Mosavie M, Pearson M et al. Bacterial genotypic and patient risk factors for adverse outcomes in Escherichia coli bloodstream infections: a prospective molecular epidemiological study. J Antimicrob Chemother 2022; 77:1753–1761 [View Article] [PubMed]
     [Google Scholar]
  12. European Committee on Antimicrobial Susceptibility Testing EUCAST Quality Control. EUCAST 2022 https://www.eucast.org/ast_of_bacteria/quality_control
     [Google Scholar]
  13. Bolger AM, Lohse M, Usadel B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 2014; 30:2114–2120 [View Article] [PubMed]
     [Google Scholar]
  14. De Coster W, D’Hert S, Schultz DT, Cruts M, Van Broeckhoven C. NanoPack: visualizing and processing long-read sequencing data. Bioinformatics 2018; 34:2666–2669 [View Article] [PubMed]
     [Google Scholar]
  15. Vaser R, Šikić M. Time- and memory-efficient genome assembly with Raven. Nat Comput Sci 2021; 1:332–336 [View Article]
     [Google Scholar]
  16. Wick RR, Judd LM, Gorrie CL, Holt KE. Unicycler: resolving bacterial genome assemblies from short and long sequencing reads. PLoS Comput Biol 2017; 13:e1005595 [View Article] [PubMed]
     [Google Scholar]
  17. Wick RR, Holt KE. Polypolish: short-read polishing of long-read bacterial genome assemblies. PLoS Comput Biol 2022; 18:e1009802 [View Article] [PubMed]
     [Google Scholar]
  18. Zimin AV, Marçais G, Puiu D, Roberts M, Salzberg SL et al. The MaSuRCA genome assembler. Bioinformatics 2013; 29:2669–2677 [View Article] [PubMed]
     [Google Scholar]
  19. Wick RR, Judd LM, Holt KE. Assembling the perfect bacterial genome using Oxford Nanopore and Illumina sequencing. PLoS Comput Biol 2023; 19:e1010905 [View Article] [PubMed]
     [Google Scholar]
  20. Seemann T. Prokka: rapid prokaryotic genome annotation. Bioinformatics 2014; 30:2068–2069 [View Article] [PubMed]
     [Google Scholar]
  21. Siguier P, Perochon J, Lestrade L, Mahillon J, Chandler M. ISfinder: the reference centre for bacterial insertion sequences. Nucleic Acids Res 2006; 34:D32–D36 [View Article] [PubMed]
     [Google Scholar]
  22. Li H. Minimap2: pairwise alignment for nucleotide sequences. Bioinformatics 2018; 34:3094–3100 [View Article] [PubMed]
     [Google Scholar]
  23. Danecek P, Bonfield JK, Liddle J, Marshall J, Ohan V et al. Twelve years of SAMtools and BCFtools. Gigascience 2021; 10:giab008 [View Article] [PubMed]
     [Google Scholar]
  24. Barrick JE, Colburn G, Deatherage DE, Traverse CC, Strand MD et al. Identifying structural variation in haploid microbial genomes from short-read resequencing data using breseq. BMC Genomics 2014; 15:1039 [View Article] [PubMed]
     [Google Scholar]
  25. Marçais G, Delcher AL, Phillippy AM, Coston R, Salzberg SL et al. MUMmer4: A fast and versatile genome alignment system. PLoS Comput Biol 2018; 14:e1005944 [View Article] [PubMed]
     [Google Scholar]
  26. Cingolani P, Platts A, Wang LL, Coon M, Nguyen T et al. A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff. Fly 2012; 6:80–92 [View Article]
     [Google Scholar]
  27. Carver T, Harris SR, Berriman M, Parkhill J, McQuillan JA. Artemis: an integrated platform for visualization and analysis of high-throughput sequence-based experimental data. Bioinformatics 2012; 28:464–469 [View Article] [PubMed]
     [Google Scholar]
  28. Alikhan N-F, Petty NK, Ben Zakour NL, Beatson SA. BLAST Ring Image Generator (BRIG): simple prokaryote genome comparisons. BMC Genomics 2011; 12:402 [View Article] [PubMed]
     [Google Scholar]
  29. Siguier P, Gourbeyre E, Varani A, Ton-Hoang B, Chandler M. Everyman’s guide to bacterial insertion sequences. Microbiol Spectr 2015; 3: [View Article] [PubMed]
     [Google Scholar]
  30. Rahn A, Beis K, Naismith JH, Whitfield C. A novel outer membrane protein, Wzi, is involved in surface assembly of the Escherichia coli K30 group 1 capsule. J Bacteriol 2003; 185:5882–5890 [View Article] [PubMed]
     [Google Scholar]
  31. ISO ISO 20776-1:2019 susceptibility testing of infectious agents and evaluation of performance of antimicrobial susceptibility test devices — part 1: broth micro-dilution reference method for testing the in vitro activity of antimicrobial agents against rapidly growing aerobic bacteria involved in infectious diseases. ISO 2019 https://www.iso.org/standard/70464.html
     [Google Scholar]
  32. EUCAST Broth microdilution - EUCAST reading guide v4.0 2022 https://www.eucast.org/fileadmin/src/media/PDFs/EUCAST_files/Disk_test_documents/2022_manuals/Reading_guide_BMD_v_4.0_2022.pdf
     [Google Scholar]
  33. Vallée M, Harding C, Hall J, Aldridge PD, TAN A. Exploring the in situ evolution of Nitrofurantoin resistance in clinically derived uropathogenic Escherichia coli isolates. J Antimicrob Chemother 2022dkac398
     [Google Scholar]
  34. Mowbray C, Tan A, Vallée M, Fisher H, Chadwick T et al. Multidrug-resistant uro-associated Escherichia coli populations and recurrent urinary tract infections in patients performing clean intermittent self-catheterisation. Eur Urol Open Sci 2022; 37:90–98 [View Article] [PubMed]
     [Google Scholar]
  35. Grindley NDF, Sherratt DJ. Sequence analysis at IS1 insertion sites: models for transposition. Cold Spring Harb Symp Quant Biol 1979; 43 Pt 2:1257–1261 [View Article] [PubMed]
     [Google Scholar]
  36. Roemhild R, Linkevicius M, Andersson DI. Molecular mechanisms of collateral sensitivity to the antibiotic nitrofurantoin. PLoS Biol 2020; 18:e3000612 [View Article] [PubMed]
     [Google Scholar]
  37. EUCAST Clinical breakpoints and dosing of antibiotics (v13.0) n.d https://www.eucast.org/clinical_breakpoints
     [Google Scholar]
  38. Muller AE, Verhaegh EM, Harbarth S, Mouton JW, Huttner A. Nitrofurantoin’s efficacy and safety as prophylaxis for urinary tract infections: a systematic review of the literature and meta-analysis of controlled trials. Clin Microbiol Infect 2017; 23:355–362 [View Article] [PubMed]
     [Google Scholar]
  39. Huttner A, Wijma RA, Stewardson AJ, Olearo F, Von Dach E et al. The pharmacokinetics of nitrofurantoin in healthy female volunteers: a randomized crossover study. J Antimicrob Chemother 2019; 74:1656–1661 [View Article] [PubMed]
     [Google Scholar]
  40. Pereira C, Warsi OM, Andersson DI. Pervasive selection for clinically relevant resistance and media adaptive mutations at very low antibiotic concentrations. Mol Biol Evol 2023; 40:msad010 [View Article] [PubMed]
     [Google Scholar]
  41. Sandegren L, Lindqvist A, Kahlmeter G, Andersson DI. Nitrofurantoin resistance mechanism and fitness cost in Escherichia coli. J Antimicrob Chemother 2008; 62:495–503 [View Article] [PubMed]
     [Google Scholar]
  42. Kram KE, Finkel SE. Culture volume and vessel affect long-term survival, mutation frequency, and oxidative stress of Escherichia coli. Appl Environ Microbiol 2014; 80:1732–1738 [View Article] [PubMed]
     [Google Scholar]
  43. Liu X, Painter RE, Enesa K, Holmes D, Whyte G et al. High-throughput screening of antibiotic-resistant bacteria in picodroplets. Lab Chip 2016; 16:1636–1643 [View Article] [PubMed]
     [Google Scholar]
  44. Reif HJ, Saedler H. IS1 is involved in deletion formation in the gal region of E. coli K12. Mol Gen Genet 1975; 137:17–28 [View Article] [PubMed]
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/mgen/10.1099/mgen.0.001102
Loading
IS1-related large-scale deletion of chromosomal regions harbouring the oxygen-insensitive nitroreductase gene nfsB causes nitrofurantoin heteroresistance in Escherichia coli
M Gen 9, 001102 (2023); https://doi.org/10.1099/mgen.0.001102
/content/journal/mgen/10.1099/mgen.0.001102
/content/journal/mgen/10.1099/mgen.0.001102
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error