1887

Abstract

Carbapenem-resistant (CRKP) has become a serious threat to global public health. Colistin is regarded as the last-resort antibiotic for CRKP infections, but colistin resistance among CRKP is increasingly being reported, making clinical treatment for CRKP infections more difficult.

The molecular mechanisms of colistin resistance in spp. under the pressure of colistin have not been fully investigated.

We aimed to investigate the phenotypic and genetic variation in two colistin-susceptible spp. strains under selective pressure of colistin.

One hundred microlitres of overnight cultures of the CRKP clinical strain CRKP12-130 and of ATCC 700603 was spread on five Mueller-Hinton Agar (MHA) plates with colistin concentrations of 2, 4, 8, 16 and 32 µg ml, and growth of colonies was observed for five consecutive days. Colonies collected from plates were passaged daily for 10 days on MHA plates without colistin and susceptibility testing of colistin was performed by broth microdilution. Thirty-four colistin-resistant strains randomly selected were submitted to whole genome sequencing (WGS). Transcriptional levels of genes involved in colistin resistance (, , , , , , and ) were measured by quantitative real-time PCR.

A total of 114 and 119 colistin-resistant colonies were obtained from CRKP12-130 and ATCC 700603 in this study, among which 16 and 18 colonies were submitted to WGS, respectively. Among these 34 sequenced isolates, mutation in (13/16, 81.25 %) was the main genetic factor mediating colistin resistance in strains from CRKP12-130, while for strains from ATCC 700603, mutation associated with (8/18, 44.44 %) was found to be the commonest. Mutation of led to a significant increase in the MIC for colistin (from 64 to >128 µg ml), and a novel mutation C28R in was first reported in this study.

Colistin-resistant spp. could be easily selected under pressure of different concentrations of colistin. Mutations of , , and genes were the main mechanisms leading to chromosomally mediated colistin resistance in spp.

Funding
This study was supported by the:
  • Shanghai Sailing Program (Award 19YF1405400)
    • Principle Award Recipient: JiachunSu
  • National Natural Science Foundation of China (Award 81903673)
    • Principle Award Recipient: JiachunSu
Loading

Article metrics loading...

/content/journal/jmm/10.1099/jmm.0.001647
2023-01-10
2024-03-29
Loading full text...

Full text loading...

References

  1. Ernst CM, Braxton JR, Rodriguez-Osorio CA, Zagieboylo AP, Li L et al. Adaptive evolution of virulence and persistence in carbapenem-resistant Klebsiella pneumoniae. Nat Med 2020; 26:705–711 [View Article] [PubMed]
    [Google Scholar]
  2. China Antimicrobial Surveillance Network; 2022 http://www.chinets.com accessed 6 July 2022
  3. Wang M, Earley M, Chen L, Hanson BM, Yu Y et al. Clinical outcomes and bacterial characteristics of carbapenem-resistant Klebsiella pneumoniae complex among patients from different global regions (CRACKLE-2): a prospective, multicentre, cohort study. Lancet Infect Dis 2022; 22:401–412 [View Article] [PubMed]
    [Google Scholar]
  4. Doi Y. Treatment options for carbapenem-resistant Gram-negative bacterial infections. Clin Infect Dis 2019; 69:S565–S575 [View Article] [PubMed]
    [Google Scholar]
  5. SENTRY Antimicrobial Surveillance Program; 2022 https://sentry-mvp.jmilabs.com/app/sentry-public/heatmap accessed 6 July 2022
  6. Zhang H, Srinivas S, Xu Y, Wei W, Feng Y. Genetic and biochemical mechanisms for bacterial lipid A modifiers associated with polymyxin resistance. Trends Biochem Sci 2019; 44:973–988 [View Article] [PubMed]
    [Google Scholar]
  7. Poirel L, Jayol A, Nordmann P. Polymyxins: antibacterial activity, susceptibility testing, and resistance mechanisms encoded by plasmids or chromosomes. Clin Microbiol Rev 2017; 30:557–596 [View Article] [PubMed]
    [Google Scholar]
  8. Magalhães C, Lima M, Trieu-Cuot P, Ferreira P. To give or not to give antibiotics is not the only question. Lancet Infect Dis 2021; 21:e191–e201 [View Article] [PubMed]
    [Google Scholar]
  9. Su J, Li D, Guo Q, Guo Y, Zheng Y et al. In vitro bactericidal activity of trimethoprim-sulfamethoxazole/colistin combination against carbapenem-resistant Klebsiella pneumoniae clinical isolates. Microb Drug Resist 2019; 25:152–156 [View Article] [PubMed]
    [Google Scholar]
  10. Elliott AG, Ganesamoorthy D, Coin L, Cooper MA, Cao MD. Complete genome sequence of Klebsiella quasipneumoniae subsp. similipneumoniae strain ATCC 700603. Genome Announc 2016; 4:e00438-16 [View Article] [PubMed]
    [Google Scholar]
  11. CLSI Performance Standards forAntimicrobial Susceptibility Testing. 32nd ed. CLSI supplement M100 Clinical and Laboratory Standards Institute; 2022
    [Google Scholar]
  12. Zhang H, Wang J, Zhou W, Yang M, Wang R et al. Risk factors and prognosis of carbapenem-resistant Klebsiella pneumoniae Infections in respiratory intensive care unit: a retrospective study. Infect Drug Resist 2021; 14:3297–3305 [View Article] [PubMed]
    [Google Scholar]
  13. Maynard M, Drusano GL, Vicchiarelli M, Liu W, Myrick J et al. Polymyxin B Pharmacodynamics in the hollow-fiber infection model: what you see may not be what you get. Antimicrob Agents Chemother 2021; 65:e0185320 [View Article] [PubMed]
    [Google Scholar]
  14. Binsker U, Käsbohrer A, Hammerl JA. Global colistin use: a review of the emergence of resistant Enterobacterales and the impact on their genetic basis. FEMS Microbiol Rev 2022; 46:fuab049 [View Article] [PubMed]
    [Google Scholar]
  15. Matheeussen V, Xavier BB, Mermans I, De Weerdt A, Lammens C et al. Emergence of colistin resistance during treatment of recurrent pneumonia caused by carbapenemase producing Klebsiella pneumoniae. Diagn Microbiol Infect Dis 2019; 94:407–409 [View Article] [PubMed]
    [Google Scholar]
  16. Yang T-Y, Wang S-F, Lin J-E, Griffith BTS, Lian S-H et al. Contributions of insertion sequences conferring colistin resistance in Klebsiella pneumoniae. Int J Antimicrob Agents 2020; 55:105894 [View Article] [PubMed]
    [Google Scholar]
  17. Cheng Y-H, Lin T-L, Pan Y-J, Wang Y-P, Lin Y-T et al. Colistin resistance mechanisms in Klebsiella pneumoniae strains from Taiwan. Antimicrob Agents Chemother 2015; 59:2909–2913 [View Article] [PubMed]
    [Google Scholar]
  18. Olaitan AO, Diene SM, Kempf M, Berrazeg M, Bakour S et al. Worldwide emergence of colistin resistance in Klebsiella pneumoniae from healthy humans and patients in Lao PDR, Thailand, Israel, Nigeria and France owing to inactivation of the PhoP/PhoQ regulator mgrB: an epidemiological and molecular study. Int J Antimicrob Agents 2014; 44:500–507 [View Article] [PubMed]
    [Google Scholar]
  19. Wright MS, Suzuki Y, Jones MB, Marshall SH, Rudin SD et al. Genomic and transcriptomic analyses of colistin-resistant clinical isolates of Klebsiella pneumoniae reveal multiple pathways of resistance. Antimicrob Agents Chemother 2015; 59:536–543 [View Article] [PubMed]
    [Google Scholar]
  20. Cannatelli A, Giani T, D’Andrea MM, Di Pilato V, Arena F et al. MgrB inactivation is a common mechanism of colistin resistance in KPC-producing Klebsiella pneumoniae of clinical origin. Antimicrob Agents Chemother 2014; 58:5696–5703 [View Article] [PubMed]
    [Google Scholar]
  21. Torres DA, Seth-Smith HMB, Joosse N, Lang C, Dubuis O et al. Colistin resistance in Gram-negative bacteria analysed by five phenotypic assays and inference of the underlying genomic mechanisms. BMC Microbiol 2021; 21:321 [View Article]
    [Google Scholar]
  22. Nordmann P, Jayol A, Poirel L. Rapid detection of polymyxin resistance in Enterobacteriaceae. Emerg Infect Dis 2016; 22:1038–1043 [View Article]
    [Google Scholar]
  23. Lippa AM, Goulian M. Perturbation of the oxidizing environment of the periplasm stimulates the PhoQ/PhoP system in Escherichia coli. J Bacteriol 2012; 194:1457–1463 [View Article]
    [Google Scholar]
  24. Cannatelli A, Santos-Lopez A, Giani T, Gonzalez-Zorn B, Rossolini GM. Polymyxin resistance caused by mgrB inactivation is not associated with significant biological cost in Klebsiella pneumoniae. Antimicrob Agents Chemother 2015; 59:2898–2900 [View Article]
    [Google Scholar]
  25. Arena F, Henrici De Angelis L, Cannatelli A, Di Pilato V, Amorese M et al. Colistin resistance caused by inactivation of the MgrB regulator is not associated with decreased virulence of sequence type 258 KPC carbapenemase-producing Klebsiella pneumoniae. Antimicrob Agents Chemother 2016; 60:2509–2512 [View Article]
    [Google Scholar]
  26. Jayol A, Nordmann P, Brink A, Poirel L. Heteroresistance to colistin in Klebsiella pneumoniae associated with alterations in the PhoPQ regulatory system. Antimicrob Agents Chemother 2015; 59:2780–2784 [View Article] [PubMed]
    [Google Scholar]
  27. Aravind L, Anantharaman V, Balaji S, Babu MM, Iyer LM. The many faces of the helix-turn-helix domain: transcription regulation and beyond. FEMS Microbiol Rev 2005; 29:231–262 [View Article] [PubMed]
    [Google Scholar]
  28. Lu HF, Tsai YC, Li LH, Lin YT, Yang TC. Role of AzoR, a LysR-type transcriptional regulator, in SmeVWX pump-mediated antibiotic resistance in Stenotrophomonas maltophilia. J Antimicrob Chemother 2021; 76:2285–2293 [View Article] [PubMed]
    [Google Scholar]
  29. Cannatelli A, D’Andrea MM, Giani T, Di Pilato V, Arena F et al. In vivo emergence of colistin resistance in Klebsiella pneumoniae producing KPC-type carbapenemases mediated by insertional inactivation of the PhoQ/PhoP mgrB regulator. Antimicrob Agents Chemother 2013; 57:5521–5526 [View Article] [PubMed]
    [Google Scholar]
  30. Jayol A, Poirel L, Brink A, Villegas M-V, Yilmaz M et al. Resistance to colistin associated with a single amino acid change in protein PmrB among Klebsiella pneumoniae isolates of worldwide origin. Antimicrob Agents Chemother 2014; 58:4762–4766 [View Article] [PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/jmm.0.001647
Loading
/content/journal/jmm/10.1099/jmm.0.001647
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF
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