Article

Letter to the Editor

Ann Lab Med 2021; 41(4): 436-438

Published online July 1, 2021 https://doi.org/10.3343/alm.2021.41.4.436

Copyright © Korean Society for Laboratory Medicine.

First Identification of IMP-1 Metallo-β-Lactamase in Delftia tsuruhatensis Strain CRS1243 Isolated From a Clinical Specimen

Sun-Mi Cho, M.D.1 , Seong Geun Hong, M.D., Ph.D.1 , Yangsoon Lee, M.D., Ph.D.2 , Wonkeun Song, M.D., Ph.D.3 , Dongeun Yong, M.D., Ph.D.4 , Seok Hoon Jeong, M.D., Ph.D.4 , Kyungwon Lee, M.D., Ph.D.4 , and Yunsop Chong, Ph.D.4

1Department of Laboratory Medicine, CHA Bundang Medical Center, CHA University, Seongnam, Korea; 2Department of Laboratory Medicine, Hanyang University College of Medicine, Seoul, Korea; 3Department of Laboratory Medicine, Hallym University College of Medicine, Seoul, Korea; 4Department of Laboratory Medicine and Research Institute of Bacterial Resistance, Yonsei University College of Medicine, Seoul, Korea

Correspondence to: Seong Geun Hong, M.D., Ph.D.
Department of Laboratory Medicine, CHA Bundang Medical Center, CHA University, 59 Yatap-ro, Bundang-gu, Seongnam 13496, Korea
Tel: +82-31-780-5463, Fax: +82-31-780-5476
E-mail: hlseo@cha.ac.kr

Received: June 26, 2020; Revised: September 23, 2020; Accepted: January 4, 2021

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Dear Editor,

Delftia tsuruhatensis is a gram-negative bacillus that was first isolated from activated sludge collected from a domestic wastewater treatment plant in Tsuruhata, Japan [1]. The bacteria are motile, slightly curved, and short rods. The species is closely related to and often misidentified as Delftia acidovorans (formerly Comamonas acidovorans) in biochemical tests because of their shared characteristics [1, 2]. However, the original D. tsuruhatensis isolate cannot utilize D-mannitol, whereas most D. acidovorans strains can [3]. We present the first report of an IMP-1 metallo-β-lactamase (MBL)-producing D. tsuruhatensis strain that was isolated from a clinical specimen. The Institutional Review Board of the CHA Bundang Medical Center, Seongnam, Korea, approved this study (approval number: 2020-04-011-001) and waived the need for informed consent.

D. tsuruhatensis is typically susceptible to carbapenem [2, 4]; however, we isolated a carbapenem-resistant strain from a 65-yr-old man diagnosed as having stomach cancer who underwent total gastrectomy. On post-operative day 16, bacterial culture was performed because of onset of fever. A Delftia species (strain CRS1243) was isolated from the surgical drainage fluid. The isolate was identified as D. acidovorans with 99% probability using Vitek 2 GN cards (bioMerieux, Marcy-l’Etoile, France) and with a score of 2.40 using the MALDI Biotyper (Bruker Daltonics, Billerica, MA, USA). However, as the isolated bacteria were unable to metabolize D-mannitol, we performed 16S rRNA gene sequencing and identified the strain as D. tsuruhatensis (100% identity with GenBank accession number HQ731453.1).

The isolate was suspected to produce carbapenemases based on routine antimicrobial susceptibility testing using Vitek AST N212 cards (bioMerieux). Antimicrobial susceptibility was determined according to the CLSI Minimum Inhibitory Concentration (MIC) Interpretive Standards for other non-Enterobacteriaceae using the broth microdilution method [5, 6]. The isolate was not susceptible to amikacin (MIC > 64 μg/mL), levofloxacin (8 μg/mL), ceftazidime (>16 μg/mL), cefepime (>32 μg/mL), ceftriaxone (>32 μg/mL), and meropenem (16 μg/mL), but it was susceptible to piperacillin-tazobactam (16 μg/mL) and minocycline (≤0.5 μg/mL). The modified Hodge test result was positive, and in carbapenemase inhibition testing (Rosco Diag-nostica A/S, Taastrup, Denmark), we observed enlarged inhibition zones when using disks containing meropenem supplemented with dipicolinic acid, indicating production of MBL. An expanded inhibition zone was also detected with disks containing meropenem supplemented with cloxacillin, but not with those containing boronic acid. Moreover, the cefoxitin-Hodge test result was negative. Based on these results, we concluded that this isolate was negative for AmpC β-lactamase activity.

Multiplex PCR was performed to detect the specific MBL. The specific primer pairs for the detection of the New Delhi MBL and Sao Paulo MBL were designed in this study and those for the detection of Verona integron-encoded MBL, imipenemase (IMP), Seoul IMP, and German IMP were selected from a previous report (Table 1). The sequence of the bacterial integron carrying the MBL-encoding gene was identified using primer walking (Table 1). We found that D. tsuruhatensis CRS1243 harbored a gene encoding blaIMP-1 within a class 1 integron located on a Tn402-like transposon. Between the 5'-conserved segment and the tni module, the gene cassettes included orfE, aac(6')-31-like, orfE, orfE, aac(6')-II, aacA7, blaIMP-1, aacA7, aac(6')-II, and qacE2 (Fig. 1). The nucleotide sequence of the class 1 integron has been deposited in GenBank under accession number KC170993. While a class 3 integron has been previously identified in a D. tsuruhatensis strain, the gene cassettes were not evaluated functionally [12]. To determine the location of the class 1 integron, we performed plasmid extraction and conjugation [13]. However, we could not identify the band corresponding to the plasmid DNA, and the carbapenem resistance was not transferred to sodium azide-resistant Escherichia coli J53.

Table 1 . Primers used for the PCR analysis of metallo-β-lactamase genes and primer walking

PrimerSequence (5´ → 3´)References
NDM-F1GCC CAA TAT TAT GCA CCC GGThis study
NDM-RCGG AAT GGC TCA TCA CGA TCThis study
SPM-NFTGC GGG AGC GCC ATT GTC TGThis study
SPM-NRTTC CAC CCG TGC CGT CCA AAThis study
VIM-FGAT GGT GTT TGG TCG CAT A7
VIM-RCGA ATG CGC AGC ACC AG7
IMP-FGGA ATA GAG TGG CTT AAY TCT C7
IMP-RCCA AAC YAC TAS GTT ATC T7
SIM-FTAC AAG GGA TTC GGC ATC G7
SIM-RTAA TGG CCT GTT CCC ATG TG7
GIM-FTCG ACA CAC CTT GGT CTG AA7
GIM-RAAC TTC CAA CTT TGC CAT GC7
INT1-5CSGGC ATC CAA GCA GCA AGC8
intl1-1FACA TGC GTG TAA ATC ATC GTC G9
attI-RCTT TGT TTT AGG GCG ACT GCThis study
aac6-F1GCT CGT TGA GAT GCT GTT CAThis study
aacA7-RGAA GCA GCG TAC TTG AGC AAThis study
IMP-1RCCT TTA ACC GCC TGC TCT AAT G10
IMP14AGG CGT GCT GCT GCA ACG ACT TGT11
qacEd1-RTGA GCC CCA TAC CTA CAA AGC9
qacE2-RATT TGA GTG TCA GCG ACA GGThis study
TniR-1GTG TTC GGT ATT TTT GCC GCThis study
TniR-2GTA ATC CCG AGT TCT TCG CAThis study
TniQ-1TGT GGT TTC GAC TTC TTC GCThis study
TniQ-2GAC CAG AAT AGC TTT GCC TGThis study
TniQ-1MTGT GGT TTC GAC TGC TAC GCThis study
TniB-1GGA AAT GGA GCA ACT GGC TThis study
TniB-2TTT CCA ACT GGT CAT CGG AGThis study
TniB-1MAGA AAT GGA ACA ACT GGC GThis study
TniA-1TAG AGC GCT GGC TCA CAT TThis study
TniA-2GGA TGT GGT CGA TGA CAA AGThis study

Figure 1. Schematic representation of the 8,846-bp partial DNA sequence of the class 1 integron containing blaIMP-1 within a Tn402-like module in Delftia tsuruhatensis CRS1243.

Cases of human infection with D. tsuruhatensis are rare [2]. In 2011, D. tsuruhatensis was identified as the etiologic agent of a human catheter-related infection; since then, it has also been associated with other human infections [2]. Recently, Fenollar, et al. [4] identified D. tsuruhatensis as an emerging opportunistic pathogen that should be considered as a cause of infection in patients with underlying disease and those using intravascular devices. The D. tsuruhatensis strain identified in this study was isolated twice in five days from pure cultures from surgical drainage fluid. Treatment with ciprofloxacin, an empirical antibiotic was started; however, piperacillin-tazobactam was administered after confirmation of the antibiotic susceptibility results. The patient’s fever subsided one day after initiation of the treatment.

To the best of our knowledge, this is the first report of IMP-1 MBL production from a D. tsuruhatensis strain. Our findings suggest that clinical microbiologists need to be aware of D. tsuruhatensis as a potential cause of opportunistic infections. We note that the blaIMP-1 gene linked to a mobile element might spread among Delftia or other bacterial species.

Hong SG conceptualized and designed the study and coordinated the drafting of the manuscript. Hong SG performed the microbiologic test and PCR work. Hong SG, Cho SM, and Lee Y performed data analysis and wrote the manuscript. Song W, Yong D, Jeong SH, Lee K, and Chong Y supervised the study, reviewed and commented on the manuscript, and approved the final draft. All authors read and approved the final manuscript.

The authors declare no conflicts of interest.

  1. Shigematsu T, Yumihara K, Ueda Y, Numaguchi M, Morimura S, Kida K. Delftia tsuruhatensis sp. nov., a terephthalate-assimilating bacterium isolated from activated sludge. Int J Syst Evol Microbiol 2003; 53: 1479-83.
    Pubmed CrossRef
  2. Preiswerk B, Ullrich S, Speich R, Bloemberg GV, Hombach M. Case report human infection with Delftia tsuruhatensis isolated from a central venous catheter. J Med Microbiol 2011; 60: 246-8.
    Pubmed CrossRef
  3. Jorgensen NO, Brandt KK, Nybroe O, Hansen M. Delftia lacustris sp. nov., a peptidoglycan-degrading bacterium from fresh water, and emended description of Delftia tsuruhatensis as a peptidoglycan-degrading bacterium. Int J Syst Evol Microbiol 2009; 59: 2195-9.
    Pubmed CrossRef
  4. Ranc A, Dubourg G, Fournier PE, Raoult D, Fenollar F. Delftia tsuruhatensis, an emergent opportunistic healthcare-associated pathogen. Emerg Infect Dis 2018; 24: 594-6.
    Pubmed KoreaMed CrossRef
  5. CLSI. Performance standards for antimicrobial susceptibility testing; approved standard. CLSI M100-S28. Wayne, PA, Clinical and Laboratory Standards Institut. 2018.
  6. CLSI. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically; approved standard. CLSI M7-A11. Wayne, PA, Clinical and Laboratory Standards Institut. 2018.
  7. Ellington MJ, Kistler J, Livermore DM, Woodford N. Multiplex PCR for rapid detection of genes encoding acquired metallo-beta-lactamases. J Antimicrob Chemother 2007; 59: 321-2.
    Pubmed CrossRef
  8. Sandvang D, Aarestrup FM, Jensen LB. Characterisation of integrons and antibiotic resistance genes in Danish multiresistant Salmonella enterica Typhimurium DT104. FEMS Microbiol Lett 1997; 157: 177-81.
    Pubmed CrossRef
  9. Zhao WH, Hu ZQ. IMP-type metallo-β-lactamases in Gram-negative bacilli: distribution, phylogeny, and association with integrons. Crit Rev Microbiol 2011; 37: 214-26.
    Pubmed CrossRef
  10. Jeong SH, Bae IK, Park KO, An YJ, Sohn SG, Jang SJ, Sung KH, Yang KS, Lee K, Young D, Lee SH. Outbreaks of imipenem-resistant Acinetobacter baumannii producing carbapenemases in Korea. J Microbiol 2006; 44: 423-31.
  11. Chu YW, Afzal-Shah M, Houang ET, Palepou MI, Lyon DJ, Woodford N, Livermore DM. IMP-4, a novel metallo-beta-lactamase from nosocomial Acinetobacter spp. collected in Hong Kong between 1994 and 1998. Antimicrob Agents Chemother 2001; 45: 710-4.
    Pubmed KoreaMed CrossRef
  12. Xu H, Davies J, Miao V. Molecular characterization of class 3 integrons from Delftia spp. J Bacteriol 2007; 189: 6276-83.
    Pubmed KoreaMed CrossRef
  13. Walsh TR, Weeks J, Livermore DM, Toleman MA. Dissemination of NDM-1 positive bacteria in the New Delhi environment and its implications for human health: an environmental point prevalence study. Lancet Infect Dis 2011; 11: 355-62.
    CrossRef