Research Article
24 October 2017

Antimicrobial Activity of Ceftazidime-Avibactam Tested against Multidrug-Resistant Enterobacteriaceae and Pseudomonas aeruginosa Isolates from U.S. Medical Centers, 2013 to 2016

ABSTRACT

The in vitro activity of ceftazidime-avibactam and many comparator agents was determined against various resistant subsets of organisms selected among 36,380 Enterobacteriaceae and 7,868 Pseudomonas aeruginosa isolates. The isolates were consecutively collected from 94 U.S. hospitals, and all isolates were tested for susceptibility by reference broth microdilution methods in a central monitoring laboratory (JMI Laboratories). Enterobacteriaceae isolates resistant to carbapenems (CRE) and/or ceftazidime-avibactam (MIC ≥ 16 μg/ml) were evaluated for the presence of genes encoding extended-spectrum β-lactamases and carbapenemases. Ceftazidime-avibactam inhibited >99.9% of all Enterobacteriaceae at the susceptible breakpoint of ≤8 μg/ml and was active against multidrug-resistant (MDR; n = 2,953; MIC50/90, 0.25/1 μg/ml; 99.2% susceptible), extensively drug-resistant (XDR; n = 448; MIC50/90, 0.5/2 μg/ml; 97.8% susceptible), and CRE (n = 513; MIC50/90, 0.5/2 μg/ml; 97.5% susceptible) isolates. Only 82.2% of MDR Enterobacteriaceae (n = 2,953) and 64.2% of ceftriaxone-nonsusceptible Klebsiella pneumoniae (n = 1,063) isolates were meropenem susceptible. Among Enterobacter cloacae (22.2% ceftazidime nonsusceptible), 99.8% of the isolates, including 99.3% of the ceftazidime-nonsusceptible isolates, were ceftazidime-avibactam susceptible. Only 23 of 36,380 Enterobacteriaceae (0.06%) isolates were ceftazidime-avibactam nonsusceptible, including 9 metallo-β-lactamase producers and 2 KPC-producing strains with porin alteration; the remaining 12 strains showed negative results for all β-lactamases tested. Ceftazidime-avibactam showed potent activity against P. aeruginosa (MIC50/90, 2/4 μg/ml; 97.1% susceptible), including MDR (MIC50/90, 4/16 μg/ml; 86.5% susceptible) isolates, and inhibited 71.8% of isolates nonsusceptible to meropenem, piperacillin-tazobactam, and ceftazidime (n = 628). In summary, ceftazidime-avibactam demonstrated potent activity against a large collection (n = 44,248) of contemporary Gram-negative bacilli isolated from U.S. patients, including organisms resistant to most currently available agents, such as CRE and meropenem-nonsusceptible P. aeruginosa.

INTRODUCTION

Widespread antimicrobial resistance and limited treatment options are increasing challenges for clinicians managing Gram-negative infections in hospital settings (1, 2). Infection control practices and new antimicrobial development have targeted mainly Gram-positive infections for many years, contributing to increasing incidences of Gram-negative infections and more antimicrobial resistance among these organisms (37). As a group, aerobic Gram-negative bacilli are the most common cause of nosocomial infections and the most common cause of infection in the intensive care unit; these organisms are very efficient at upregulating and acquiring antimicrobial resistance genes (2, 8, 9). Furthermore, infections caused by multidrug-resistant (MDR) Gram-negative organisms cause significant morbidity and mortality, longer hospitalizations, and increased costs compared with infections caused by susceptible organisms, and the Gram-negative infections require promptly introducing effective antimicrobial therapy (10).
β-Lactamase production remains the major factor in rising antimicrobial resistance in Gram-negative bacteria, and the most clinically relevant β-lactamases produced by Gram-negative organisms isolated from U.S. medical centers are the extended-spectrum β-lactamases (ESBLs) CTX-M and SHV and Klebsiella pneumoniae carbapenemase (KPC) (2, 11). The wide dissemination of these β-lactamases has left clinicians and patients with very few treatment options for infections caused by MDR Enterobacteriaceae (12, 13).
Ceftazidime-avibactam is a combination agent consisting of the β-lactamase inhibitor avibactam and the broad-spectrum cephalosporin ceftazidime (14, 15). Avibactam is a member of a novel class of non-β-lactam β-lactamase inhibitors, the diazabicyclooctanes (DBO), and acts as a reversible, covalent inhibitor. Compared to available inhibitors for clinical use, DBOs are more potent, have a broader spectrum, and have a different mechanism of action. A unique feature of avibactam compared to earlier β-lactamase inhibitors is that avibactam binds reversibly to β-lactamases, allowing for recyclization and inhibition of additional β-lactamase molecules. Avibactam effectively inactivates class A (ESBLs and KPC), class C (AmpC), and some class D (such as OXA-48) β-lactamases (16).
Ceftazidime-avibactam was approved by the U.S. Food and Drug Administration (FDA) and by the European Medicine Agency (EMA) to treat complicated intra-abdominal infection in combination with metronidazole, as well as complicated urinary tract infections, including pyelonephritis (17, 18). Ceftazidime-avibactam is also approved to treat nosocomial pneumonia in Europe and has been studied in pediatric patients (NCT01893346) (19). As part of the International Network for Optimal Resistance Monitoring (INFORM) surveillance program, we evaluated the activity of ceftazidime-avibactam against a large collection of contemporary (2013 to 2016) MDR Gram-negative organisms causing infections in patients from U.S. medical centers.

RESULTS

Ceftazidime-avibactam inhibited >99.9% of all Enterobacteriaceae isolates (n = 36,380) at the susceptible breakpoint of ≤8 μg/ml (Tables 1 and 2) and was highly active against MDR isolates (n = 2,953; MIC50/90, 0.25/1 μg/ml; 99.2% susceptible), extensively drug-resistant (XDR) isolates (n = 448; MIC50/90, 0.5/2 μg/ml; 97.8% susceptible), and carbapenem-resistant (CRE) isolates (n = 513; MIC50/90, 0.5/2 μg/ml; 97.5% susceptible; Tables 1 and 2) isolates. Amikacin (MIC50/90, 2/4 μg/ml; 99.2% susceptible according to the Clinical and Laboratory Standards Institute [CLSI] criteria), tigecycline (MIC50/90, 0.25/1 μg/ml; 98.0% susceptible according to the FDA), and meropenem (MIC50/90, ≤0.06/≤0.06 μg/ml; 98.5% susceptible according to the CLSI) were also very active against the entire collection of Enterobacteriaceae isolates, but these antimicrobial agents exhibited limited activity against MDR, XDR, and CRE isolates (Table 2).
TABLE 1
TABLE 1 Antimicrobial activity of ceftazidime-avibactam tested against antimicrobial resistant Enterobacteriaceae and P. aeruginosa from U.S. hospitals (2013 to 2016)a
Organism (no. of isolates)No. of isolates and cumulative % at various MICs (μg/ml)bMIC (μg/ml)
≤0.0150.030.060.120.250.512481632>32MIC50MIC90
Enterobacteriaceae (36,380)1,3344,42811,13912,3424,8151,631448142631513190.120.25
3.715.846.580.493.698.199.399.799.999.9>99.9>99.9100.0
    CRE (513)165133576137138592014090.52
3.14.16.613.528.355.081.993.497.397.598.298.2100.0
    MDR (2,953)98217428597587525296116531313190.251
3.310.725.245.465.383.093.197.098.899.299.799.7100.0
    XDR (448)169202553126117442354060.52
3.65.610.015.627.555.681.791.596.797.898.798.7100.0
E. coli (12,942)1,0241,4634,5974,5501,0661863613310030.060.25
7.919.254.789.998.199.699.899.9>99.9>99.9>99.9>99.9100.0
    Ceftriaxone-NS (1,750)100632477364441083312310030.120.25
5.79.323.465.590.997.098.999.699.899.899.899.8100.0
    MEM-NS (27)11358411000030.25>32
3.77.418.537.066.781.585.288.988.988.988.988.9100.0
K. pneumoniae (7,511)1373642,5232,950957364137581701030.120.25
1.86.740.379.592.397.198.999.799.999.9>99.9>99.9100.0
    MEM-NS (382)1557215611199481601030.52
3.95.27.112.627.256.382.294.899.099.099.299.2100.0
    Ceftriaxone-NS (1,063)392367259242229127561701030.251
3.75.812.136.559.380.892.898.099.699.699.799.7100.0
    Colistin-NS (205)94224930363213900010.252
4.46.317.141.055.673.288.895.199.599.599.599.5100.0
K. oxytoca (1,902)89382370917474145110.120.25
0.45.348.685.995.098.999.699.999.9100.0
Proteus mirabilis (2,818)421,5681,1028314330110010.030.06
1.557.196.299.299.799.899.999.999.9>99.9>99.9>99.9100.0
E. cloacae (3,740)33262151,5941,18447716433803120.250.5
0.91.67.349.981.694.498.799.699.899.899.999.9100.0
    CAZ-NS (831)13374422934514630803120.51
1.61.92.88.135.677.194.798.399.399.399.699.8100.0
E. aerogenes (1,384)2947352605256751045010.120.25
2.15.530.974.693.198.699.399.699.999.9100.0
    CAZ-NS (279)107176411349945010.250.5
3.66.112.235.175.693.296.497.899.699.6100.0
Morganella morganii (1,059)23704891204025921010.060.12
0.235.181.392.696.498.899.699.899.999.9100.0
Citrobacter koseri (777)34943732056010010.060.12
4.416.564.590.998.699.999.9100.0
Citrobacter freundii (1,026)121517147322598232610.120.5
1.22.619.365.487.396.999.199.399.9100.0
Serratia marcescens (1,715)4497206682333151220.250.5
0.23.145.184.097.699.499.799.899.9100.0
Proteus vulgaris (418)318618538240.060.12
0.745.289.598.699.0100.0
Providencia spp. (872)516418423114974141918950.120.5
0.619.440.567.084.192.594.296.398.499.4100.0
P. aeruginosa (7,868)1413882,7712,7791,137426134434924
1.86.741.977.391.797.198.899.4100.0
    CAZ-NS (1,204)117933173442161334349416
0.10.20.78.534.863.481.392.495.9100.0
    MEM-NS (1,471)481213824852821073745416
0.30.89.035.068.087.294.496.9100.0
    P-T-NS (1,497)21151083604673361253944416
0.10.21.28.432.563.786.194.597.1100.0
    CAZ, MEM, and P-T-NS (628)221061741491013541832
3.520.448.171.887.993.5100.0
    MDR (1,562)8181293844983141224247416
0.51.79.934.566.486.594.397.0100.0
    XDR (717)10132126190194933644832
0.10.10.34.722.348.875.988.893.9100.0
a
Abbreviations: CRE, carbapenem-resistant Enterobacteriaceae; MDR, multidrug resistant; XDR, extensively drug resistant; NS, nonsusceptible; MEM, meropenem; CAZ, ceftazidime; P-T, piperacillin-tazobactam.
b
For each entry in column 1, there are two rows of data. The first row indicates the number of strains, and the second row indicates the cumulative percentage. Column subheadings specify the various MICs in μg/ml.
TABLE 2
TABLE 2 Activity of ceftazidime-avibactam and comparator antimicrobial agents when tested against Enterobacteriaceae, P. aeruginosa, and selected resistant subsetsa
Organism category and antimicrobial agent (no. of isolates tested)MIC (μg/ml)CLSIbEUCAST
MIC50MIC90%S%R%S%R
Enterobacteriaceae
    All isolates (36,380)
        Ceftazidime-avibactam0.120.2599.90.1*99.90.1
        Ceftriaxone≤0.06>885.313.685.313.6
        Ceftazidime0.25888.89.986.211.2
        Cefepime≤0.12291.26.789.87.7
        Piperacillin-tazobactam21692.74.189.87.3
        Meropenem≤0.06≤0.0698.51.398.70.7
        Levofloxacin≤0.12>482.615.678.718.8
        Gentamicin≤1291.27.690.28.8
        Amikacin2499.20.298.40.8
        Tigecycline0.25198.00.1*92.72.0
        Colistin≤0.5>878.221.8
    MDR (2,953)c
        Ceftazidime-avibactam0.25199.20.8*99.20.8
        Ceftriaxone>8>819.777.219.777.2
        Ceftazidime32>3229.663.922.170.4
        Cefepime16>1637.451.132.056.8
        Piperacillin-tazobactam16>6452.729.845.247.3
        Meropenem≤0.06882.215.384.78.9
        Levofloxacin>4>420.670.611.284.8
        Gentamicin>8>838.250.834.561.8
        Amikacin41691.12.284.48.9
        Tigecycline0.5488.90.3*76.911.1
        Colistin≤0.5>860.739.3
    XDR (448)d
        Ceftazidime-avibactam0.5297.82.2*97.82.2
        Ceftriaxone>8>82.097.32.097.3
        Ceftazidime>32>325.491.52.994.6
        Cefepime>16>1610.579.96.485.8
        Piperacillin-tazobactam>64>647.183.76.592.9
        Meropenem8>821.272.527.548.2
        Levofloxacin>4>48.784.82.296.4
        Gentamicin8>827.050.023.473.0
        Amikacin163260.29.646.539.8
        Tigecycline0.5490.00.2*81.010.0
        Colistin≤0.5>861.338.7
    CRE (513)e
        Ceftazidime-avibactam0.5297.52.5*97.52.5
        Ceftriaxone>8>82.197.52.197.5
        Ceftazidime>32>324.393.02.395.7
        Cefepime>16>168.477.93.287.1
        Piperacillin-tazobactam>64>643.191.22.796.9
        Meropenem>8>82.789.710.352.4
        Levofloxacin>4>423.472.915.081.3
        Gentamicin8>849.533.944.450.5
        Amikacin83268.27.051.531.8
        Tigecycline0.5198.80.0*90.31.2
        Colistin≤0.5>879.120.9
    Ceftriaxone-nonsusceptible K. pneumoniae (1,063)f
        Ceftazidime-avibactam0.25199.60.4*99.60.4
        Ceftriaxone>8>80.097.50.097.5
        Ceftazidime>32>329.084.53.491.0
        Cefepime>16>1612.374.48.981.6
        Piperacillin-tazobactam32>6444.744.331.855.3
        Meropenem≤0.06>864.232.867.222.3
        Levofloxacin>4>435.158.624.167.9
        Gentamicin8>846.545.342.153.5
        Amikacin43281.04.270.319.0
        Tigecycline0.5198.80.0*91.21.2
        Colistin≤0.5488.911.1
    Colistin-nonsusceptible K. pneumoniae (205)
        Ceftazidime-avibactam0.25299.50.5*99.50.5
        Ceftriaxone>8>842.955.642.955.6
        Ceftazidime32>3242.057.140.058.0
        Cefepime16>1646.751.146.751.1
        Piperacillin-tazobactam16>6451.245.941.048.8
        Meropenem≤0.06>862.437.662.431.2
        Levofloxacin>4>446.850.242.054.1
        Gentamicin≤1>869.825.965.930.2
        Amikacin23277.67.366.822.4
        Tigecycline0.5299.50.0*89.30.5
        Colistin>8>80.0100.0
    Ceftazidime-nonsusceptible (≥8 μg/ml) Enterobacter cloacae (831)g
        Ceftazidime-avibactam0.5199.30.7*99.30.7
        Ceftriaxone>8>80.099.40.099.4
        Ceftazidime>32>320.095.80.0100.0
        Cefepime2>1665.118.343.124.1
        Piperacillin-tazobactam64>6423.835.916.676.2
        Meropenem≤0.060.2593.55.194.91.3
        Levofloxacin≤0.12>482.312.975.520.6
        Gentamicin≤1>883.512.882.216.5
        Amikacin1499.30.698.00.7
        Tigecycline0.25298.30.0*89.21.7
        Colistin≤0.5>884.615.4
P. aeruginosa
    All isolates (7,868)
        Ceftazidime-avibactam2497.12.9*97.12.9
        Ceftazidime23284.710.984.715.3
        Cefepime21685.65.285.614.4
        Piperacillin-tazobactam46481.09.481.019.0
        Meropenem0.5881.312.881.36.8
        Levofloxacin0.5>474.518.665.334.7
        Gentamicin2887.08.487.013.0
        Amikacin4896.51.991.83.5
        Colistin1299.60.499.60.4
    MDR (1,562)
        Ceftazidime-avibactam41686.513.5*86.513.5
        Ceftazidime16>3243.641.643.656.4
        Cefepime16>1641.723.041.758.3
        Piperacillin-tazobactam64>6431.236.231.268.8
        Meropenem8>820.058.220.032.4
        Levofloxacin>4>418.765.09.590.5
        Gentamicin4>855.432.355.444.6
        Amikacin83287.17.673.912.9
        Colistin1299.20.899.20.8
    XDR (717)
        Ceftazidime-avibactam83275.924.1*75.924.1
        Ceftazidime32>3221.560.021.578.5
        Cefepime16>1617.338.117.382.7
        Piperacillin-tazobactam>64>647.752.67.792.3
        Meropenem>8>83.179.63.152.2
        Levofloxacin>4>45.378.42.297.8
        Gentamicin8>839.647.139.660.4
        Amikacin8>3280.811.664.219.2
        Colistin1299.30.799.30.7
    Ceftazidime, meropenem, and piperacillin-tazobactam-nonsusceptible (628)
        Ceftazidime-avibactam83271.828.2*71.828.2
        Ceftazidime32>320.076.00.0100.0
        Cefepime16>1618.341.918.381.7
        Piperacillin-tazobactam>64>640.065.10.0100.0
        Meropenem>8>80.081.20.050.6
        Levofloxacin>4>421.966.213.686.4
        Gentamicin4>854.037.454.046.0
        Amikacin8>3283.010.771.717.0
        Colistin1299.50.599.50.5
a
CLSI and EUCAST criteria are as published previously (33, 34).
b
*, breakpoints from the FDA package insert (17, 35).
c
MDR organisms included the following: Citrobacter freundii (n = 59), C. koseri (n = 1), Enterobacter aerogenes (n = 42), E. cloacae (n = 119), E. cloacae species complex (n = 153), Escherichia coli (n = 797), Hafnia alvei (n = 3), Klebsiella oxytoca (n = 55), K. pneumoniae (n = 793), Morganella morganii (n = 174), Proteus mirabilis (n = 426), P. vulgaris (n = 5), P. vulgaris group (n = 1), Providencia rettgeri (n = 17), P. stuartii (n = 191), Raoultella ornithinolytica (n = 1), Serratia liquefaciens (n = 1), S. marcescens (n = 112), Raoultella sp. (n = 2), and Serratia sp. (n = 1).
d
XDR organisms included the following: Citrobacter freundii (n = 9), Enterobacter aerogenes (n = 1), E. cloacae (n = 44), Escherichia coli (n = 12), Klebsiella oxytoca (n = 8), K. pneumoniae (n = 297), Morganella morganii (n = 17), Proteus mirabilis (n = 19), Providencia stuartii (n = 18), Raoultella ornithinolytica (n = 1), Serratia marcescens (n = 21), and Raoultella sp. (n = 1).
e
The results of the molecular characterization of these isolates are shown in Table 3. CRE organisms included the following: Citrobacter freundii (n = 6), Enterobacter aerogenes (n = 13), E. cloacae (n = 5), E. cloacae species complex (n = 50), Escherichia coli (n = 24), Klebsiella oxytoca (n = 16), K. pneumoniae (n = 368), Proteus mirabilis (n = 4), Providencia stuartii (n = 2), Raoultella ornithinolytica (n = 1), Serratia marcescens (n = 22), and Raoultella sp. (n = 2).
f
According to CLSI criteria; ≥2 μg/ml (33).
g
According to CLSI criteria; ≥8 μg/ml (33).
The most active compound tested against MDR and XDR Enterobacteriaceae isolates was ceftazidime-avibactam (99.2 and 97.8% susceptible, respectively, per FDA and European Committee on Antimicrobial Susceptibility Testing [EUCAST] criteria), followed by amikacin (91.1 and 60.2% susceptible, respectively, per CLSI criteria and 84.4 and 46.5% susceptible, respectively, per EUCAST criteria), tigecycline (88.9 and 90.0% susceptible, respectively, per FDA criteria and 76.9 and 81.0% susceptible, respectively per EUCAST criteria), and meropenem (82.2 and 21.2% susceptible, respectively, per CLSI criteria and 84.7 and 27.5% susceptible, respectively, per EUCAST criteria; Table 2). Among CRE, 97.5% of isolates were susceptible to ceftazidime-avibactam (FDA and EUCAST criteria; MIC50/90, 0.5/2 μg/ml), 98.8/90.3% of isolates were susceptible (FDA/EUCAST criteria) to tigecycline, and 68.2/51.5% were susceptible (CLSI/EUCAST) to amikacin. Furthermore, only 79.1% of CRE isolates were susceptible to colistin per EUCAST criteria (≤2 μg/ml; Table 2).
β-Lactamase gene screening results for the 513 CRE isolates are summarized in Table 3. The most common acquired carbapenemase gene observed among CRE isolates was blaKPC (435 isolates; 84.8%), mainly blaKPC-3 (220 isolates; 42.9% of total) and blaKPC-2 (129 isolates; 25.1%), and the KPC-producing isolates were very susceptible to ceftazidime-avibactam (MIC50/90, 0.5/2 μg/ml; data not shown). Resistance to ceftazidime-avibactam (MIC value of 16 μg/ml) was observed in two KPC-producing isolates, which also displayed porin alterations.
TABLE 3
TABLE 3 Combinations detected among 513 CRE isolates
Enzyme(s)No. of isolatesOrganism (no. of isolates)
Citrobacter freundiiEnterobacter aerogenesEscherichia coliEnterobacter cloacaeKlebsiella oxytocaKlebsiella pneumoniaeProteus mirabilisProvidencia stuartiiRaoultella spp.aSerratia marcescens
KPC-212922101029814
KPC-3220314291415838
KPC-4431
KPC-1711
KPC-like7979
KPC-2, VIM-411
KPC-17, NDM-111
OXA-23211
OXA-4811
SME-455
NDM-14211
NDM-911
IMP-like, NMC-A11
IMP-27b11b
IMP-6411
VIM-111
Total positive4515317461634121317
Negative resultsc62110790272105
Total51361324551636842322
a
These included two Raoultella ornithinolytica isolates and one Raoultella sp. isolate.
b
The ceftazidime-avibactam MIC for this P. mirabilis isolate is 4 μg/ml.
c
No carbapenemase or ESBL genes were detected.
A metallo-β-lactamase (MBL) gene was only observed in 11 Enterobacteriaceae isolates (0.03%; Table 3), and these isolates exhibited decreased susceptibility to ceftazidime-avibactam and all β-lactam compounds tested (data not shown). Only two isolates carrying an MBL gene were susceptible to ceftazidime-avibactam, one P. mirabilis isolate and one Providencia rettgeri isolate with ceftazidime-avibactam MIC values of 2 and 4 μg/ml, respectively, both carrying a blaIMP-27. Other acquired carbapenemase genes found among CRE isolates included blaOXA-48, blaOXA-232, blaSME-4, and blaNMC-A (Table 3).
Overall, only 23 of 36,380 Enterobacteriaceae (0.06%) were ceftazidime-avibactam nonsusceptible, including 6 NDM-producing strains (2 Escherichia coli, 2 K. pneumoniae, and 1 Enterobacter cloacae with ceftazidime-avibactam MIC values of >32 μg/ml; Table 3), 1 VIM-4 producing K. pneumoniae (ceftazidime-avibactam MIC of >32 μg/ml), 1 IMP-64-producing isolate (Proteus mirabilis with ceftazidime-avibactam MIC of >32 μg/ml), 1 IMP-27-producing Morganella morganii (ceftazidime-avibactam MIC of 16 μg/ml), 2 KPC-producing isolates (K. pneumoniae and E. cloacae with ceftazidime-avibactam MIC of 16 μg/ml), and 12 isolates (4 E. cloacae, 1 E. aerogenes, 5 Providencia stuartii, and 2 Serratia marcescens) with negative results for all β-lactamases tested (data not shown).
Resistance to meropenem (MIC ≥ 2 μg/ml; nonsusceptible per CLSI criteria) was observed in 27 (0.2%) E. coli and 382 (5.1%) K. pneumoniae isolates, and ceftazidime-avibactam retained activity against 88.9 and 99.0%, respectively, of these isolates (Tables 1 and 2). The ceftazidime-avibactam-nonsusceptible isolates were 3 E. coli (all NDM-producing) and 4 K. pneumoniae (2 NDM-1-, 1 VIM-4-, and 1 KPC-2-producing strains with porin alterations). Tigecycline was also active against meropenem-nonsusceptible E. coli (100.0%) and K. pneumoniae (99.0% susceptible per FDA criteria), whereas colistin was active against 100.0% of the E. coli strains but only 79.6% of K. pneumoniae isolates nonsusceptible to meropenem (Table 2). All other compounds demonstrated very limited activity against these organisms (Table 2). Ceftazidime-avibactam was also active against colistin-resistant K. pneumoniae (n = 205; MIC50/90, 0.25/2 μg/ml; 99.5% susceptible) and ceftazidime-nonsusceptible E. cloacae (n = 831; MIC50/90, 0.5/1 μg/ml; 99.3% susceptible; Table 2).
Ceftazidime-avibactam exhibited potent activity against Pseudomonas aeruginosa (n = 7,868; MIC50/90, 2/4 μg/ml; 97.1% susceptible), including most MDR (n = 1,562; MIC50/90, 4/16 μg/ml; 86.5% susceptible) and XDR (n = 717; MIC50/90, 8/32 μg/ml; 75.9% susceptible) isolates. Furthermore, ceftazidime-avibactam retained in vitro activity against P. aeruginosa isolates nonsusceptible to meropenem (MIC50/90, 4/16 μg/ml; 87.2% susceptible), piperacillin-tazobactam (MIC50/90, 4/16 μg/ml; 86.1% susceptible), or ceftazidime (MIC50/90, 4/16 μg/ml; 81.3% susceptible), as well as isolates nonsusceptible to meropenem, piperacillin-tazobactam and ceftazidime (MIC50/90, 8/32 μg/ml; 71.8% susceptible; Table 1). These P. aeruginosa-resistant subsets exhibited high rates of resistance to all comparator agents tested, except amikacin and colistin. Amikacin was active against 87.1% of MDR, 80.8% of XDR, and 83.0% of isolates nonsusceptible to ceftazidime, meropenem, and piperacillin-tazobactam at the CLSI susceptible breakpoint of ≤16 μg/ml; however, >99% of isolates from these resistant subsets remained susceptible to colistin (Table 2).

DISCUSSION

The shortage of active compounds and a deficiency of convincing clinical data have led to controversy about antimicrobial treatment choices for infections caused by MDR and XDR Gram-negative bacilli (20, 21). For many years, the treatment of these infections, especially those caused by CRE, relied on various combination regimens with fragile clinical data to support their use. In terms of in vitro activity, colistin, tigecycline, fosfomycin, and some aminoglycosides are among the few agents that may remain active against these organisms; however, all of these compounds have important spectrum deficiencies that prevent their use for empirical treatment of life-threatening infections (21, 22).
Ceftazidime-avibactam approval brought to the market a significant option for treating MDR Gram-negative infections (15, 16, 23). The addition of avibactam restores the activity of ceftazidime against Gram-negative bacilli that acquired β-lactam resistance through expression of the Ambler class A ESBLs, chromosomal or mobile class C β-lactamases, serine carbapenemases, or some class D β-lactamases. The results of this investigation highlight the main problems of antimicrobial resistance in U.S. medical centers and indicate that ceftazidime-avibactam represents a valuable option for the treatment of infections caused by antimicrobial-resistant Gram-negative bacilli, including MDR and XDR strains.
More than 44,000 contemporary Gram-negative bacteria were evaluated, including a large number of isolates resistant to multiple antimicrobial agents, and ceftazidime-avibactam was active against 99.9% of Enterobacteriaceae isolates, including 97.5% of CRE, 99.2% of MDR, and 97.8% of XDR isolates. Our results also showed that the KPC family of enzymes represent the main cause of carbapenem resistance among Enterobacteriaceae in U.S. hospitals, and these carbapenemases appear to be very well inhibited by avibactam. Ceftazidime-avibactam-nonsusceptible Enterobacteriaceae were limited to MBL-producing strains (9 isolates; 0.02% of the collection), 2 KPC-producing strains with porin alterations, and 12 strains (0.03% of the collection) with no ceftazidime-hydrolyzing enzyme detected. Resistance to ceftazidime-avibactam in these 12 isolates is probably due to porin alteration and/or hyperexpression of efflux pumps; however, further characterization of these isolates is necessary to confirm this hypothesis and exclude the production of novel β-lactamases that are not effectively inhibited by avibactam (24).
The results of this investigation also confirmed the potent in vitro activity and broad coverage of this compound against P. aeruginosa. Ceftazidime-avibactam inhibited 97.1% of P. aeruginosa isolates at the susceptible breakpoint of ≤8 μg/ml, including 86.5% of MDR, 75.9% of XDR, and 71.8% of isolates resistant to ceftazidime, meropenem, and piperacillin-tazobactam. It is important to note that the ceftazidime-avibactam MIC values were generally higher among the resistant subsets of P. aeruginosa (MIC50, 4 to 8 μg/ml) compared to the wild-type (ceftazidime-susceptible) group (MIC50, 2 μg/ml; data not shown), possibly indicating the expression of multiple mechanisms of resistance (25). Among non-β-lactam compounds tested, only amikacin (80.8 to 87.1% susceptible according to CLSI criteria) and colistin (99.2 to 99.5% susceptible according to CLSI and EUCAST criteria) showed reasonable in vitro activity against these resistant subsets of P. aeruginosa. Further characterization of these ceftazidime-resistant isolates is currently being performed to elucidate the resistance mechanism.
In summary, ceftazidime-avibactam demonstrated potent in vitro activity and broad antimicrobial spectrum against a large collection (n = 44,248) of contemporary Gram-negative bacilli isolated from patients in 94 U.S. hospitals during a 4-year period (2013 to 2016). Our results coupled with pharmacokinetic/pharmacodynamic, clinical, and safety results published to date (2628) indicate that ceftazidime-avibactam represents a valuable treatment option for infections caused by Enterobacteriaceae and P. aeruginosa, including those caused by organisms resistant to most antimicrobial agents currently available.

MATERIALS AND METHODS

Bacterial isolates.

A total of 36,380 Enterobacteriaceae and 7,868 P. aeruginosa isolates were collected from 94 medical centers among 39 states from all 9 U.S. Census divisions in 2013 to 2016 as part of the INFORM program (29). These isolates were collected from patients with pneumonia (n = 11,666; 26.4%), bloodstream infections (n = 6,110; 13.8%), skin and skin structure infections (n = 9,728; 22.0%), urinary tract infections (n = 12,688; 28.7%), intra-abdominal infections (n = 2,038; 4.6%), and other infection types (n = 2,018; 4.6%) according to defined protocols (30). Only isolates determined to be significant by local criteria as the reported probable cause of infection was included in the program. Species identification was confirmed by standard biochemical tests and using a MALDI Biotyper (Bruker Daltonics, Billerica, MA) according to the manufacturer's instructions, where necessary.

Resistant subsets.

Isolates were categorized as MDR, XDR, or pan drug-resistant (PDR) according to criteria published by Magiorakos et al. (31), which defines MDR as nonsusceptible to ≥1 agent in ≥3 antimicrobial classes, XDR as nonsusceptible to ≥1 agent in all but ≤2 antimicrobial classes, and PDR as nonsusceptible (CLSI criteria) to all antimicrobial classes tested. The antimicrobial classes and drug representatives used in the analysis for Enterobacteriaceae were: broad-spectrum cephalosporins (ceftriaxone, ceftazidime, and cefepime), carbapenems (imipenem, meropenem, and doripenem), broad-spectrum penicillin combined with a β-lactamase-inhibitor (piperacillin-tazobactam), fluoroquinolones (ciprofloxacin and levofloxacin), aminoglycosides (gentamicin, tobramycin, and amikacin), glycylcyclines (tigecycline), and the polymyxins (colistin [EUCAST criteria]). Those used in the analysis for P. aeruginosa included antipseudomonal cephalosporins (ceftazidime and cefepime), carbapenems (imipenem, meropenem, and doripenem), broad-spectrum penicillins combined with a β-lactamase-inhibitor (piperacillin-tazobactam), fluoroquinolones (ciprofloxacin and levofloxacin), aminoglycosides (gentamicin, tobramycin, and amikacin), and polymyxins (colistin). In addition, CRE was defined as resistant (MIC, ≥4 μg/ml [CLSI]) to imipenem (imipenem was not applied to Proteus mirabilis or to indole-positive Proteeae), meropenem, or doripenem.

Susceptibility testing.

Broth microdilution test methods conducted according to the CLSI were performed to determine the antimicrobial susceptibility of ceftazidime-avibactam (inhibitor at fixed concentration of 4 μg/ml) and comparator agents (32). Concurrent quality control (QC) testing was performed to ensure proper test conditions and procedures. QC strains included E. coli ATCC 25922 and 35218 and P. aeruginosa ATCC 27853. All QC results were within published ranges. CLSI (33) and EUCAST (34) susceptibility interpretive criteria were used to determine susceptibility/resistance rates for comparator agents. Furthermore, FDA and EUCAST breakpoint criteria were applied for ceftazidime-avibactam when testing Enterobacteriaceae and P. aeruginosa, susceptible at ≤8 μg/ml and resistant at ≥16 μg/ml (17).

Screening for β-lactamases.

Enterobacteriaceae isolates displaying elevated ceftazidime-avibactam MIC values (≥16 μg/ml) and all CRE isolates were tested for β-lactamase-encoding genes using the microarray-based assay Check-MDR CT101 kit (Check-Points, Wageningen, Netherlands) and/or next-generation sequencing (NGS). The Check-MDR CT101 assay was performed according to the manufacturer's instructions. This kit has the capabilities to detect CTX-M groups 1, 2, 8+25, and 9; TEM wild type (WT) and ESBL SHV WT; and ESBL, ACC, ACT/MIR, CMYII, DHA, FOX, KPC, and NDM-1 (11). Total genomic DNA was extracted using the fully automated Thermo Scientific KingFisher Flex Magnetic Particle Processor (Cleveland, OH). To perform NGS, DNA extracts were quantified using the Qubit High Sensitivity DS-DNA assay (Invitrogen/Thermo Fisher, Inc.) and normalized to 0.2 ng/μl. A total of 1 ng of high-quality genomic DNA was used as input material for library construction using the Nextera XT DNA library preparation kit (Illumina, San Diego, CA). Libraries were normalized using the bead-based normalization procedure (Illumina) and sequenced on MiSeq. Fastq files generated were assembled using SPAdes Assembler and subjected to a proprietary software (JMI Laboratories) for screening of β-lactamase genes.

ACKNOWLEDGMENTS

We thank all participants of the International Network for Optimal Resistance Monitoring (INFORM) program for providing bacterial isolates.
This study was supported by Allergan. Allergan was involved in the design and decision to present these results, and JMI Laboratories received compensation fees for services in relation to preparing the manuscript. Allergan had no involvement in the collection, analysis, and interpretation of data.
JMI Laboratories contracted to perform services in 2016 for Achaogen, Actelion, Allecra Therapeutics, Allergan, AmpliPhi Biosciences; API; Astellas Pharma; AstraZeneca; Basilea Pharmaceutica; Bayer AG; BD; Biomodels; Cardeas Pharma Corp.; CEM-102 Pharma; Cempra; Cidara Therapeutics, Inc.; CorMedix; CSA Biotech; Cutanea Life Sciences, Inc.; Debiopharm Group; Dipexium Pharmaceuticals, Inc.; Duke; Entasis Therapeutics, Inc.; Fortress Biotech; Fox Chase Chemical Diversity Center, Inc.; Geom Therapeutics, Inc.; GSK; Laboratory Specialists, Inc.; Medpace; Melinta Therapeutics, Inc.; Merck & Co., Inc.; Micromyx; MicuRx Pharmaceuticals, Inc.; Motif Bio; N8 Medical, Inc.; Nabriva Therapeutics, Inc.; Nexcida Therapeutics, Inc.; Novartis; Paratek Pharmaceuticals, Inc.; Pfizer; Polyphor; Rempex; Scynexis; Shionogi; Spero Therapeutics; Symbal Therapeutics; Synlogic; TenNor Therapeutics; TGV Therapeutics; The Medicines Company; Theravance Biopharma; ThermoFisher Scientific; VenatoRx Pharmaceuticals, Inc.; and Wockhardt, Zavante Therapeutics, Inc. There are no speakers' bureaus or stock options to declare.

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Information & Contributors

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Published In

cover image Antimicrobial Agents and Chemotherapy
Antimicrobial Agents and Chemotherapy
Volume 61Number 11November 2017
eLocator: 10.1128/aac.01045-17

History

Received: 18 May 2017
Returned for modification: 19 June 2017
Accepted: 14 August 2017
Published online: 24 October 2017

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Keywords

  1. antimicrobial resistance
  2. carbapenem-resistant Enterobacteriaceae
  3. beta-lactamases

Contributors

Authors

Helio S. Sader
JMI Laboratories, North Liberty, Iowa, USA
Mariana Castanheira
JMI Laboratories, North Liberty, Iowa, USA
Dee Shortridge
JMI Laboratories, North Liberty, Iowa, USA
Rodrigo E. Mendes
JMI Laboratories, North Liberty, Iowa, USA
Robert K. Flamm
JMI Laboratories, North Liberty, Iowa, USA

Notes

Address correspondence to Helio S. Sader, [email protected].

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