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Gonzalez, McMullen, Wallace, Crotty, Ritchie, and Burnham: Susceptibility of Ceftolozane-Tazobactam and Ceftazidime-Avibactam Against a Collection of β-Lactam-Resistant Gram-Negative Bacteria
Letter to the Editor
Clinical Microbiology

Annals of Laboratory Medicine 2017; 37(2): 174-176.

Published online: 20 December 2016

DOI: https://doi.org/10.3343/alm.2017.37.2.174

Susceptibility of Ceftolozane-Tazobactam and Ceftazidime-Avibactam Against a Collection of β-Lactam-Resistant Gram-Negative Bacteria

Mark D. Gonzalez, Ph.D.1, 2, Allison R. McMullen, Ph.D.1, Meghan A. Wallace, B.S.1, Matthew P. Crotty, Pharm.D.3, 4, David J. Ritchie, Pharm.D.3, 5, Carey-Ann D. Burnham, Ph.D.1

1Department of Pathology and Immunology, Division of Laboratory and Genomic Medicine, Washington University School of Medicine, St. Louis, MO, USA.

2Department of Laboratory Medicine, Children's Healthcare of Atlanta, Atlanta, GA, USA.

3Pharmacy Department, Barnes-Jewish Hospital, St. Louis, MO, USA.

4Department of Pharmacy, Methodist Dallas Medical Center, Dallas, TX, USA.

5Pharmacy Practice Department, St. Louis College of Pharmacy, St. Louis, MO, USA.

Corresponding author: Mark D. Gonzalez. Department of Laboratory Medicine, Children's Healthcare of Atlanta, 1405 Clifton Road, NE, Atlanta, GA 30322, USA. Tel: +1-404-785-2553, Fax: +1-404-785-6342, mark.gonzalez@choa.org

Received 14 March 2016       Revised 10 August 2016       Accepted 17 November 2016

© The Korean Society for Laboratory Medicine.

(open-access, http://creativecommons.org/licenses/by-nc/4.0/):

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,
Ceftolozane-tazobactam (C/T) and ceftazidime-avibactam (CZA) were recently approved for the treatment of complicated intra-abdominal infections and complicated urinary tract infections (http://www.fda.gov/NewsEvents/Newsroom/PressAnnouncements/ucm427534.htm, http://www.fda.gov/NewsEvents/Newsroom/PressAnnouncements/ucm435629.htm, both accessed February 24, 2016). To date, only one study has simultaneously evaluated the activities of C/T and CZA in vitro against Pseudomonas aeruginosa, and few studies have evaluated the effects of these antibiotics on multi-drug resistant (MDR) gram-negative bacteria [123]. This study aimed to examine the activities of C/T and CZA against β-lactam-resistant Enterobacteriaceae and P. aeruginosa clinical isolates.
The isolates were recovered from clinical specimens at Barnes-Jewish Hospital (St. Louis, MO, USA) from September to December 2014. Specimen sources included respiratory, blood, urine, and wound samples. Isolates of Enterobacteriaceae were included if they tested non-susceptible to cefepime and/or had an extended-spectrum β-lactamase (ESBL)-producing phenotype. P. aeruginosa isolates were included if they tested non-susceptible to meropenem. We included carbapenem-resistant Enterobacteriaceae isolates recovered from August 2012 to December 2014; these strains were tested for the blaKPC and blaNDM genes by real-time PCR [45]. All isolates were negative for the blaNDM gene.
Frozen stocks of all isolates were subcultured twice consecutively on 5% sheep's blood agar (Hardy Diagnostics, Santa Maria, CA, USA) prior to antimicrobial susceptibility testing (AST). Species-level identification was confirmed by using the VITEK MS system (IVD v2.3.3, bioMérieux, Durham, NC, USA) [67]. AST was performed by using gradient diffusion (Etest, bioMérieux). In brief, a 0.5 McFarland standard suspension of each isolate was inoculated onto Mueller-Hinton agar (Hardy Diagnostics), and Etest strips were applied. Plates were incubated overnight at 35℃ in ambient air. Each day, QC strains (Escherichia coli ATCC 25922 and ATCC 35218, and P. aeruginosa ATCC 27853) were tested. C/T and CZA results and QC were interpreted by using Food and Drug Administration breakpoints. The categorical interpretation and QC ranges for all other antibiotics (BD BBL and Co., Sparks, MD, USA) were based on standardized disk diffusion criteria [8].
We evaluated 120 clinical isolates comprising 45 P. aeruginosa strains and 75 Enterobacteriaceae strains. Table 1 shows the overall β-lactam susceptibility profile. A subset of P. aeruginosa isolates (n=10, 22%), termed “β-lactam-resistant (BLR)”, were resistant to piperacillin/tazobactam, cefepime, meropenem, and imipenem.
The 50% minimum inhibitory concentration (MIC50, 1 µg/mL) and 90% minimum inhibitory concentration (MIC90, 8 µg/mL) of C/T were lower for the P. aeruginosa isolates than for the Enterobacteriaceae isolates (2 µg/mL and 32 µg/mL, respectively; Table 2). Furthermore, 87% (n=39) of all P. aeruginosa isolates and 60% (n=6) of the BLR P. aeruginosa isolates were C/T-susceptible (Table 2).
Within the Enterobacteriaceae, the C/T data showed group-dependent differences. For example, the E. coli isolates had a low MIC90 (0.5 µg/mL), whereas the Enterobacter spp. had a higher MIC90 (64 µg/mL). Overall, 56% (n=42) of all Enterobacteriaceae isolates were C/T-susceptible (Table 2).
In contrast to the C/T results, the Enterobacteriaceae had lower MIC50 (0.5 µg/mL) and MIC90 (2 µg/mL) values for CZA compared to the P. aeruginosa isolates (2 µg/mL and 16 µg/mL, respectively; Table 2). Notably, 82% (n=37) of all P. aeruginosa isolates were CZA-susceptible (Table 2), whereas 99% (n=74) of Enterobacteriaceae isolates were CZA-susceptible. An Enterobacter spp. isolate was CZA-resistant (MIC of 32 µg/mL), but was found to be negative for the blaKPC and blaNDM genes by real-time PCR.
Comparison of the P. aeruginosa C/T and CZA results showed 87% (n=39) concordance with 35 isolates testing susceptible and four isolates testing non-susceptible to both antibiotics. For the Enterobacteriaceae, there was only a 57% concordance between the C/T and CZA results; 42 isolates tested susceptible and one isolate tested non-susceptible to both antibiotics. All remaining isolates were only CZA-susceptible.
The availability of new agents with anti-gram-negative activity holds promise for treating MDR organisms. CZA provides an alternative treatment for blaKPC-positive organisms, which are otherwise treated by agents with less desirable safety or efficacy profiles. Ceftolozane is a new cephalosporin with activity against P. aeruginosa, and in combination with tazobactam shows activity against ESBL-producing Enterobacteriaceae [9]. Notably, although two of the blaKPC-positive isolates were C/T-susceptible, we would not expect C/T to be clinically effective.
In summary, we report the activities of C/T and CZA against a collection of BLR Enterobacteriaceae and P. aeruginosa isolates. Our results suggest that C/T and CZA are active against and represent possible therapeutic options for infections with BLR gram-negative bacteria.

Acknowledgments

Cubist supplied the C/T Etest strips, and Actavis supplied the CZA Etest strips. There was no funding for this study.

Notes

Authors' Disclosures of Potential Conflicts of Interest: MDG has none to declare. CAB has received research support from bioMerieux, Cepheid, and Accelerate Diagnostics.

References

1. Estabrook M, Bussell B, Clugston SL, Bush K. In vitro activity of ceftolozane-tazobactam as determined by broth dilution and agar diffusion assays against recent U.S. Escherichia coli isolates from 2010 to 2011 carrying CTX-M-type extended-spectrum β-lactamases. J Clin Microbiol. 2014; 52:4049–4052. PMID: 25143578.
2. Sader HS, Castanheira M, Mendes RE, Flamm RK, Farrell DJ, Jones RN. Ceftazidime-avibactam activity against multidrug-resistant Pseudomonas aeruginosa isolated in U.S. medical centers in 2012 and 2013. Antimicrob Agents Chemother. 2015; 59:3656–3659. PMID: 25845861.
3. Buehrle DJ, Shields RK, Chen L, Hao B, Press EG, Alkrouk A, et al. Evaluation of the in vitro activity of ceftazidime-avibactam and ceftolozane-tazobactam against meropenem-resistant Pseudomonas aeruginosa isolates. Antimicrob Agents Chemother. 2016; 60:3227–3231. PMID: 26976862.
4. Doern CD, Dunne WM Jr, Burnham CA. Detection of Klebsiella pneumoniae carbapenemase (KPC) production in non-Klebsiella pneumoniae Enterobacteriaceae isolates by use of the Phoenix, Vitek 2, and disk diffusion methods. J Clin Microbiol. 2011; 49:1143–1147. PMID: 21209164.
5. Centers for Disease Control and Prevention. Multiplex real-time PCR detection of Klebsiella pneumoniae carbapenemase (KPC) and New Delhi metallo-β-lactamase (NDM-1). 2010. http://www.cdc.gov/hai/pdfs/labsettings/kpc-ndm-protocol-2011.pdf.
6. Manji R, Bythrow M, Branda JA, Burnham CA, Ferraro MJ, Garner OB, et al. Multi-center evaluation of the Vitek® MS system for mass spectrometric identification of non-Enterobacteriaceae Gram-negative bacilli. Eur J Clin Microbiol Infect Dis. 2014; 33:337–346. PMID: 24019163.
7. Richter SS, Sercia L, Branda JA, Burnham CA, Bythrow M, Ferraro MJ, et al. Identification of Enterobacteriaceae by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using the Vitek MS system. Eur J Clin Microbiol Infect Dis. 2013; 32:1571–1578. PMID: 23818163.
8. Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing. 25th Informational supplement, M100-S25. Wayne, PA: Clinical and Laboratory Standards Institute;2015.
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Table 1

β-Lactam susceptibility profiles of the study isolates (N=120)

alm-37-174-i001
Bacterial group (N) % Susceptible*
Ceftazidime Pip-Tazo Cefepime Meropenem Ertapenem Imipenem
Pseudomonas aeruginosa (45) 73 67 71 13 NA 22
 BLR P. aeruginosa† (10) 10 0 0 0 NA 0
Enterobacteriaceae (75) 27 40 21 64 57 73
 Enterobacter spp. (17) 6 6 12 35 12 47
 Escherichia coli (29) 59 86 28 100 100 100
 Klebsiella pneumoniae (24) 8 17 8 42 42 63
  ESBL K. pneumoniae (8) 13 50 25 100 100 100
  CRE K. pneumoniae‡ (16) 6 0 0 13 13 44
Remaining isolates§ (5) 0 0 80 60 40 60
Enterobacteriaceae blaKPC status∥
 blaKPC positive (10) 0 0 10 0 10 0
 blaKPC negative** (65) 31 46 23 74 65 85

*Based on CLSI M100-S25 antibiotic disk diffusion criteria; †BLR P. aeruginosa isolates were not susceptible to piperacillin-tazobactam, cefepime, meropenem, and imipenem; ‡CRE K. pneumoniae, carbapenem-resistant K. pneumoniae isolates that either tested positive for blaKPC (n=8) or were negative for blaKPC and blaNDM (n=8) by real-time PCR; §Remaining isolates, including Citrobacter freundii complex (n=3), Klebsiella oxytoca (n=1), and Morganella morganii (n=1); ∥Enterobacteriaceae blaKPC status, all of the above Enterobacteriaceae isolates identified only by blaKPC status; blaKPC-positive, isolates that tested positive for blaKPC by real-time PCR; **blaKPC-negative, isolates that either tested negative for blaKPC and blaNDM by real-time PCR (n=20) or lacked a phenotype (n=45) that is consistent with a blaKPC-positive organism (i.e., lack of resistance to meropenem).

Abbreviations: C/T, ceftolozane-tazobactam; CZA, ceftazidime-avibactam; MIC, minimum inhibitory concentration; NA, not applicable; ESBL, extended spectrum β-lactamase; CRE, carbapenem-resistant Enterobacteriaceae.

Table 2

Ceftolozane-tazobactam (C/T) and ceftazidime-avibactam (CZA) activity against BLR gram-negative bacteria

alm-37-174-i002
Bacterial group (N) C/T MIC Range µg/mL C/T MIC50 C/T MIC90 C/T % Sus* CZA MIC Range µg/mL CZA MIC50 CZA MIC90 CZA % Sus*
Pseudomonas aeruginosa (45) 0.25–16 1 8 87 0.5–64 2 16 82
 BLR P. aeruginosa† (10) 2–16 4 8 60 2–64 8 64 50
Enterobacteriaceae (75) 0.125– ≥ 256 2 32 56 0.032–32 0.5 2 99
 Enterobacter spp. (17) 0.5–64 4 64 18 0.125–32 1 4 94
 Escherichia coli (29) 0.125–4 0.25 0.5 97 0.032–2 0.125 0.5 100
 Klebsiella pneumoniae (24) 0.25– ≥ 256 4 128 42 0.125–8 1 2 100
  ESBL K. pneumoniae (8) 0.25–4 0.25 4 88 0.125–1 0.25 1 100
  CRE K. pneumoniae‡ (16) 1– ≥ 256 8 ≥ 256 19 1–8 2 4 100
Remaining isolates§ (5) 2–16 16 16 20 0.5–2 1 2 100
Enterobacteriaceae blaKPC status∥
 blaKPC positive (10) 2–128 8 128 20 0.5–4 1 2 100
 blaKPC negative** (65) 0.125– ≥ 256 0.5 16 62 0.032–32 0.25 2 99

*% Sus, % Susceptible based on Food and Drug Administration interpretative criteria for ceftolozane-tazobactam and ceftazidime-avibactam; †BLR P. aeruginosa isolates that were not susceptible to piperacillin-tazobactam, cefepime, meropenem, and imipenem; ‡CRE K. pneumoniae, carbapenem-resistant K. pneumoniae isolates that either tested positive for blaKPC (n=8) or were negative for blaKPC and blaNDM (n=8) by real-time PCR; §Remaining isolates, including Citrobacter freundii complex (n=3), Klebsiella oxytoca (n=1), and Morganella morganii (n=1); ∥Enterobacteriaceae blaKPC status, all of the above Enterobacteriaceae isolates identified only by blaKPC status; blaKPC-positive, isolates that tested positive for blaKPC by real-time PCR; **blaKPC-negative, isolates that either tested negative for blaKPC and blaNDM by real-time PCR (n=20) or lacked a phenotype (n=45) that is consistent with a blaKPC-positive organism (i.e., lack of resistance to meropenem).

Abbreviations: C/T, ceftolozane-tazobactam; CZA, ceftazidime-avibactam; MIC, minimum inhibitory concentration; ESBL, extended spectrum β-lactamase; CRE, carbapenem-resistant Enterobacteriaceae.

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