ABSTRACT
Plazomicin and comparator agents were tested by using the CLSI reference broth microdilution method against 4,825 clinical isolates collected during 2014 and 2015 in 70 U.S. hospitals as part of the ALERT (Antimicrobial Longitudinal Evaluation and Resistance Trends) program. Plazomicin (MIC50/MIC90, 0.5/2 μg/ml) inhibited 99.2% of 4,362 Enterobacteriaceae at ≤4 μg/ml. Amikacin, gentamicin, and tobramycin inhibited 98.9%, 90.3%, and 90.3% of these isolates, respectively, by applying CLSI breakpoints. The activities of plazomicin were similar among Enterobacteriaceae species, with MIC50 values ranging from 0.25 to 1 μg/ml, with the exception of Proteus mirabilis and indole-positive Proteeae that displayed MIC50 values of 2 μg/ml. For 97 carbapenem-resistant Enterobacteriaceae (CRE), which included 87 isolates carrying blaKPC, plazomicin inhibited all but 1 isolate at ≤2 μg/ml (99.0% and 98.9%, respectively). Amikacin and gentamicin inhibited 64.9% and 56.7% of the CRE isolates at the respective CLSI breakpoints. Plazomicin inhibited 96.5 and 95.5% of the gentamicin-resistant isolates, 96.9 and 96.5% of the tobramycin-resistant isolates, and 64.3 and 90.0% of the amikacin-resistant isolates according to CLSI and EUCAST breakpoints, respectively. The activities of plazomicin against Pseudomonas aeruginosa (MIC50/MIC90, 4/16 μg/ml) and Acinetobacter species (MIC50/MIC90, 2/16 μg/ml) isolates were similar. Plazomicin was active against coagulase-negative staphylococci (MIC50/MIC90, 0.12/0.5 μg/ml) and Staphylococcus aureus (MIC50/MIC90, 0.5/0.5 μg/ml) but had limited activity against Enterococcus spp. (MIC50/MIC90, 16/64 μg/ml) and Streptococcus pneumoniae (MIC50/MIC90, 32/64 μg/ml). Plazomicin activity against the Enterobacteriaceae tested, including CRE and isolates carrying blaKPC from U.S. hospitals, supports the development plan for plazomicin to treat serious infections caused by resistant Enterobacteriaceae in patients with limited treatment options.
INTRODUCTION
The worldwide emergence of multidrug-resistant (MDR) organisms, including carbapenem-resistant Enterobacteriaceae (CRE), Pseudomonas aeruginosa, and Acinetobacter baumannii, highlights the need for new therapeutic options to treat infections (1). In the early 2000s, the Infectious Diseases Society of America recognized the urgent need for surveillance initiatives and new therapeutic options for the group of organisms known as ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, A. baumannii, P. aeruginosa, and Enterobacter species), which includes Gram-negative organisms as well as troublesome Gram-positive species (2).
New therapeutic options for Gram-positive and -negative organisms were recently approved for clinical therapy in the United States and Europe (3); however, developing new agents with broad-spectrum activity is important given the significance of adequate empirical treatment for successful patient outcomes (4, 5).
Aminoglycosides are broad-spectrum agents that have been used for several decades to treat serious infections caused by nonfastidious Gram-negative bacteria, staphylococci, enterococci, and viridans group streptococci. Aminoglycosides are also used in combination with other agents displaying synergistic activity with this class, such as β-lactams, fluoroquinolones, polymyxins, and vancomycin (3). The most common mechanisms of resistance to aminoglycosides are aminoglycoside-modifying enzymes (AMEs) that are broadly disseminated among Gram-negative and Gram-positive species and carried by mobile genetic structures that also often harbor β-lactamases and other resistance genes (6, 7).
Plazomicin is a semisynthetic aminoglycoside derived from sisomicin and contains structural modifications that make this molecule stable in the presence of most AMEs (8, 9). Plazomicin has in vitro activity against nonfastidious Gram-negative pathogens and Staphylococcus spp., including methicillin-resistant S. aureus isolates (10, 11).
In this study, we evaluated the activities of plazomicin and comparator antimicrobial agents tested against 4,825 clinical isolates collected in U.S. hospitals during 2014 and 2015. Enterobacteriaceae isolates displaying elevated carbapenem MIC values were evaluated for carbapenemase genes, and a separate analysis focused on the activities of plazomicin and comparators against these subsets.
RESULTS
Activities of plazomicin and comparator agents.Plazomicin (MIC50 and MIC90, 0.5 and 2 μg/ml, respectively) inhibited 99.2% of 4,362 Enterobacteriaceae isolates tested at ≤4 μg/ml (Table 1). For common Enterobacteriaceae species, plazomicin inhibited 99.9% of the 1,506 K. pneumoniae isolates (MIC50 and MIC90, 0.25 and 0.5 μg/ml, respectively), 99.9% of the 1,346 Escherichia coli isolates (MIC50 and MIC90, 0.5 and 1 μg/ml, respectively), and 99.4% of the 359 Klebsiella oxytoca isolates (MIC50 and MIC90, 0.5 and 0.5 μg/ml, respectively) at ≤4 μg/ml (Table 1).
Antimicrobial activity of plazomicin tested against 4,825 clinical isolates collected in 70 U.S. hospitals during 2014 to 2015
Plazomicin inhibited all 104 Enterobacter cloacae species complex isolates (here E. cloacae) (MIC50 and MIC90, 0.5 and 0.5 μg/ml, respectively) at ≤2 μg/ml and all 120 Enterobacter aerogenes (MIC50 and MIC90, 0.5 and 1 μg/ml, respectively), 159 Citrobacter freundii species complex (MIC50 and MIC90, 0.5 and 1 μg/ml, respectively), 145 Citrobacter koseri (MIC50 and MIC90, 0.25 and 0.5 μg/ml, respectively), and 107 Serratia marcescens (MIC50 and MIC90, 1 and 2 μg/ml, respectively) isolates at ≤4 μg/ml (Table 1).
Overall, plazomicin inhibited 90.7%, 91.1%, 98.4%, and 99.1% of the Morganella morganii, Providencia species, Proteus mirabilis, and Proteus vulgaris group isolates at ≤4 μg/ml, respectively (Table 1). Plazomicin MIC50 values for Morganella morganii, Proteus mirabilis, and Proteus vulgaris (2, 2, and 2 μg/ml, respectively) were identical to the MIC50 values displayed by amikacin (2, 2, and 2 μg/ml, respectively), while gentamicin (0.5, 1, and 0.5 μg/ml, respectively) and tobramycin (0.5, 0.5, and 0.5 μg/ml, respectively) (data not shown) had 2- to 4-fold-lower MIC50 values. Plazomicin displayed MIC50 values (range, 0.25 to 1 μg/ml) lower than those observed for amikacin (2 to 1 μg/ml) for all remaining Enterobacteriaceae species and were similar to those observed for gentamicin (0.5 μg/ml for all species) and tobramycin (range, 0.25 to 2 μg/ml). The MIC50 values of all 4 aminoglycosides were very similar (range, 1 to 2 μg/ml) for Providencia species isolates (data not shown).
Amikacin (MIC50 and MIC90, 1 and 4 μg/ml, respectively), gentamicin (MIC50 and MIC90, 0.5 and 4 μg/ml, respectively), and tobramycin (MIC50 and MIC90, 0.5 and 4 μg/ml, respectively) inhibited 98.9%, 90.3%, and 90.3%, respectively, of the 4,362 Enterobacteriaceae isolates tested by using the CLSI breakpoints (Table 2). Among other comparator agents, meropenem (MIC50 and MIC90, 0.03 and 0.06 μg/ml, respectively; 97.9% susceptible) and tigecycline (MIC50 and MIC90, 0.25 and 1 μg/ml, respectively; 99.0% susceptible according to U.S. Food and Drug Administration [FDA] breakpoints) were most active against the Enterobacteriaceae isolates tested (Table 2).
Activities of plazomicin and comparator agents tested against Enterobacteriaceae isolates
Plazomicin had activity against the majority of Enterobacteriaceae isolates resistant to gentamicin and tobramycin (Table 3), inhibiting 96.5% and 95.5% of the gentamicin-resistant isolates according to CLSI and EUCAST breakpoint criteria, respectively, and 96.9% and 96.5% of the tobramycin-resistant isolates according to the same criteria. Only 14 isolates were resistant to amikacin when applying the CLSI criteria, and plazomicin inhibited 9 (64.3%) of these isolates at ≤4 μg/ml. Plazomicin inhibited 90.0% of the 50 amikacin-resistant isolates identified by using the EUCAST breakpoints at ≤4 μg/ml. Five amikacin-resistant isolates had plazomicin MIC values of ≥128 μg/ml and were resistant to gentamicin and tobramycin.
Activities of plazomicin and comparator aminoglycosides against phenotypes and genotypes of Enterobacteriaceae isolates and other Gram-negative species
For 103 P. aeruginosa isolates, the plazomicin MIC50 and MIC90 values were 4 and 16 μg/ml, respectively. These values were 2-fold higher than those for amikacin and gentamicin (MIC50 and MIC90, 2 and 8 μg/ml, respectively, for both) and 8- to 16-fold higher than those for tobramycin (MIC50 and MIC90, 0.5 and 1 μg/ml, respectively) (Tables 1 and 3). Amikacin (97.1% susceptible according to CLSI breakpoints), tobramycin (94.2% susceptible according to CLSI breakpoints), and colistin (98.1% susceptible according to EUCAST breakpoints) (data not shown) were the most active comparators against P. aeruginosa isolates.
For 95 Acinetobacter species isolates, plazomicin MIC50 and MIC90 values were 4 and 16 μg/ml, respectively (Table 1). Amikacin (MIC50 and MIC90, 4 and >32 μg/ml, respectively), gentamicin (MIC50 and MIC90, 1 and >8 μg/ml, respectively), and tobramycin (MIC50 and MIC90, 1 and >8 μg/ml, respectively) (Table 3) inhibited 83.0%, 69.5%, and 77.9% of the isolates, respectively, according to CLSI breakpoint criteria. Among other comparators, only colistin (94.7% susceptible using CLSI or EUCAST criteria), ampicillin-sulbactam (72.3% using CLSI criteria), and imipenem (70.5% using both criteria) inhibited >70.0% of the isolates (data not shown).
All 72 coagulase-negative staphylococci (plazomicin MIC50 and MIC90, 0.12 and 0.5 μg/ml, respectively) were inhibited by plazomicin at ≤0.5 μg/ml, and 69 S. aureus (plazomicin MIC50 and MIC90, 0.5 and 0.5 μg/ml, respectively) (Table 1) isolates tested were inhibited by plazomicin at ≤2 μg/ml, including methicillin-resistant isolates (30 isolates tested) (Table 1). Gentamicin inhibited 72.2% and 97.1% of the respective coagulase-negative staphylococcus and S. aureus isolates tested at the CLSI breakpoints (data no shown).
Plazomicin had limited antimicrobial activity against Streptococcus pneumoniae (n = 66; MIC50 and MIC90, 32 and 64 μg/ml, respectively) and Enterococcus spp. (n = 58; MIC50 and MIC90, 16 and 64 μg/ml, respectively) (Table 1). As expected, other aminoglycosides also had activities similar to those of plazomicin against these organisms (data not shown).
Activity of plazomicin against carbapenem-resistant and carbapenemase-producing Enterobacteriaceae.A total of 97 (2.2% of Enterobacteriaceae) CRE isolates were identified in this study. Plazomicin (MIC50 and MIC90, 0.5 and 1 μg/ml, respectively) inhibited 99.0% of these isolates at an MIC of ≤2 or ≤4 μg/ml (Table 2). CRE isolates were highly resistant to comparator agents, but nearly all of them were susceptible to tigecycline (100.0% susceptible according to U.S. FDA breakpoints) (Table 2). Amikacin (MIC50 and MIC90, 16 and 32 μg/ml, respectively), gentamicin (MIC50 and MIC90, 2 and >8 μg/ml, respectively), and tobramycin (MIC50 and MIC90, >8 and >8 μg/ml, respectively) inhibited only 64.9%, 56.7%, and 13.4% of the isolates, respectively, at current CLSI breakpoints (Table 2).
Among 113 isolates displaying imipenem and/or meropenem MIC values of ≥2 μg/ml, carbapenemases were detected in 87 isolates, including 79 K. pneumoniae, 4 K. oxytoca, 2 C. freundii, 1 E. cloacae, and 1 S. marcescens isolates. All isolates carried a blaKPC variant: 56 carried blaKPC-3, 29 carried blaKPC-2, 1 carried blaKPC-4, and 1 carried blaKPC-17. These isolates were detected in all U.S. census divisions but were observed mainly in the Middle Atlantic division (52 isolates; 59.8% of the carbapenemase-producing Enterobacteriaceae [CPE]).
Plazomicin (MIC50 and MIC90, 0.25 and 1 μg/ml, respectively) inhibited 86 of the 87 (98.9%) isolates carrying blaKPC at ≤2 or ≤4 μg/ml (Tables 2 and 3). One isolate exhibited a plazomicin MIC at >128 μg/ml, was resistant to all aminoglycosides, and carried the 16S rRNA methyltransferase gene rmtF1. Amikacin (MIC50 and MIC90, 16 and 32 μg/ml, respectively), gentamicin (MIC50 and MIC90, 4 and >8 μg/ml, respectively), and tobramycin (MIC50 and MIC90, >8 and >8 μg/ml, respectively) inhibited 63.2 and 42.5%, 55.2 and 48.3%, and 11.5 and 9.2% of the respective CPE isolates according to current CLSI/EUCAST breakpoint criteria.
Carbapenemase-negative isolates included 16 K. pneumoniae isolates, 4 S. marcescens isolates, and 3 isolates of other species. Plazomicin demonstrated comparable activities against the carbapenemase-negative (MIC50 and MIC90, 0.5 and 1 μg/ml, respectively) and CPE (MIC50 and MIC90, 0.25 and 1 μg/ml, respectively) isolates, and the highest plazomicin MIC value observed was at 2 μg/ml. In contrast, the aminoglycoside comparators were more potent against the 26 carbapenemase-negative isolates than against the CPE isolates. Amikacin (MIC50 and MIC90, 4 and 16 μg/ml, respectively), gentamicin (MIC50 and MIC90, 1 and >8 μg/ml, respectively), and tobramycin (MIC50 and MIC90, 2 and >8 μg/ml, respectively) inhibited 92.3%, 69.2%, and 57.7% of the carbapenemase-negative isolates, respectively, when applying the CLSI breakpoints (Table 2).
DISCUSSION
Patients with prolonged hospitalization, including those in intensive care or long-term-care facilities, immunocompromised patients, and others with malignant conditions, often develop infections, and many of the infections are caused by MDR organisms (1). These organisms include members of the Enterobacteriaceae family, including CRE, and pandrug- and extensively drug-resistant P. aeruginosa, A. baumannii, and Gram-positive species, including E. faecium and S. aureus. The urgent need for monitoring initiatives and new potential therapeutic options for these organisms has been recognized by the medical and scientific communities, and although various new antimicrobial agents for Gram-positive infections have been approved, the number of non-β-lactam candidates for treating Gram-negative infections in late-stage development is still limited (2–5).
In this study, plazomicin displayed activity against CRE and CPE isolates detected in U.S. hospitals. The majority of the CRE isolates (and all CPE isolates) carried blaKPC, and only 1 of these isolates had a plazomicin MIC value of >4 μg/ml. CRE isolates displayed elevated rates of resistance to all β-lactams and comparator agents, including gentamicin and amikacin, which inhibited 63.2% or fewer of the blaKPC-carrying isolates at current CLSI or EUCAST breakpoints. Furthermore, plazomicin displayed activity against most isolates resistant to gentamicin and tobramycin according to CLSI or EUCAST breakpoints and inhibited >63% of the amikacin-resistant isolates.
Combination therapy is often used to treat infections caused by CRE, and this might include an aminoglycoside. The in vitro results presented here suggest that there is a potential role for this new agent in the antimicrobial armamentarium against difficult-to-treat MDR organisms. These data are supported by the results of 2 recent phase 3 clinical trials that evaluated plazomicin in complicated urinary tract infections and in serious infections due to CRE (i.e., bloodstream infections, hospital-acquired and ventilator-associated bacterial pneumonia, and complicated urinary tract infections) (ClinicalTrials.gov registration numbers NCT02486627 [https://clinicaltrials.gov/ct2/show/NCT02486627] and NCT01970371 [https://clinicaltrials.gov/ct2/show/NCT01970371]).
Plazomicin demonstrated activity against Enterobacteriaceae, Staphylococcus spp. regardless of methicillin resistance, and some P. aeruginosa and Acinetobacter species isolates. A tentative breakpoint of 4 μg/ml was applied during the plazomicin development program and thus was also applied here for the analysis of the Enterobacteriaceae population, based on pharmacokinetic/pharmacodynamic (PK/PD) data, which was supported by animal efficacy data (12). By applying this tentative breakpoint, the in vitro activity of plazomicin was similar to or greater than those of other aminoglycosides against important organism groups, which include multidrug-resistant isolates, that can pose a challenge for current antimicrobial chemotherapy options.
MATERIALS AND METHODS
Bacterial isolates.A total of 4,825 clinical isolates, including 4,362 Enterobacteriaceae, 265 Gram-positive cocci, 103 P. aeruginosa isolates, and 95 Acinetobacter spp., collected during 2014 and 2015 in 70 hospitals located in 61 U.S. cities were evaluated as part of the ALERT (Antimicrobial Longitudinal Evaluation and Resistance Trends) program. This surveillance program collects key pathogens in targeted numbers (1 per patient episode) deemed to cause urinary tract infections (1,414 isolates), bloodstream infections (1,178), pneumonia in hospitalized patients (1,125), skin and skin structure infections (566), and intra-abdominal infections (441). Other infection sources (101 isolates) were also accepted for uncommon species. Species identification was confirmed, when necessary, by matrix-assisted laser desorption ionization–time of flight mass spectrometry using the Bruker Daltonics MALDI Biotyper (Bruker Daltonics, Billerica, MA, USA), according to the manufacturer's instructions.
Antimicrobial susceptibility testing.All isolates were tested for susceptibility to plazomicin and comparator agents using the reference broth microdilution method described by the CLSI (13). Categorical interpretations for all comparator agents were found in CLSI criteria in document M100 (14), EUCAST breakpoint tables (15), and/or the FDA website (16). Quality control was performed by using E. coli ATCC 25922, S. aureus ATCC 29213, P. aeruginosa ATCC 27853, Enterococcus faecalis ATCC 29212, and S. pneumoniae ATCC 49619. All quality control MIC results were within acceptable ranges as reported in CLSI documents.
Definitions.CRE was defined as any isolate exhibiting an imipenem, meropenem, or doripenem MIC value of >2 μg/ml. Proteus mirabilis and indole-positive Proteeae were selected by using meropenem and doripenem only due to the intrinsically elevated imipenem MIC values.
Characterization of carbapenemases.Isolates nonsusceptible to imipenem or meropenem were screened by PCR followed by DNA sequencing of blaKPC, blaIMP, blaVIM, blaNDM, blaOXA-48, blaGES (blaGES-2, blaGES-4, blaGES-5, blaGES-6, and blaGES-8), blaNMC-A, blaSME, and blaIMI (17). Isolates yielding negative results for these genes were tested for less common carbapenemases, including genes encoding FRI-1, BKC-1, GIM-1/-2, SIM-1, SPM-1, KHM-1, AIM-1, BIC-1, and DIM-1 (18).
Amplicons generated were sequenced on both strands, and the nucleotide sequences obtained were analyzed by using the Lasergene software package (DNAStar, Madison, WI, USA) and compared to available sequences via an NCBI BLAST search (http://www.ncbi.nlm.nih.gov/blast/).
ACKNOWLEDGMENTS
We thank the JMI staff for their technical expertise in all stages of this study and manuscript preparation.
This study was performed by JMI Laboratories and supported by Achaogen, which included funding for services related to preparing the manuscript.
JMI Laboratories was contracted to perform services in 2017 for Achaogen; Allecra Therapeutics; Allergan; Amplyx Pharmaceuticals; Antabio; API; Astellas Pharma; AstraZeneca; Athelas; Basilea Pharmaceutica; Bayer AG; BD; Becton, Dickinson and Co.; Boston Pharmaceuticals; CEM-102 Pharma; Cempra; Cidara Therapeutics, Inc.; CorMedix; CSA Biotech; Cutanea Life Sciences, Inc.; Entasis Therapeutics, Inc.; Geom Therapeutics, Inc.; GSK; Iterum Pharma; Medpace; Melinta Therapeutics, Inc.; Merck & Co., Inc.; MicuRx Pharmaceuticals, Inc.; N8 Medical, Inc.; Nabriva Therapeutics, Inc.; NAEJA-RGM; Novartis; Paratek Pharmaceuticals, Inc.; Pfizer; Polyphor; Ra Pharma; Rempex; Riptide Bioscience, Inc.; Roche; Scynexis; Shionogi; Sinsa Labs, Inc.; Skyline Antiinfectives; Sonoran Biosciences; Spero Therapeutics; Symbiotica; Synlogic; Synthes Biomaterials; TenNor Therapeutics; Tetraphase; The Medicines Company; Theravance Biopharma; VenatoRx Pharmaceuticals, Inc.; Wockhardt; Yukon Pharma; Zai Laboratory; and Zavante Therapeutics, Inc. There are no speakers' bureaus or stock options to declare. A.W.S. and K.M.K. are employees of Achaogen and contributed with the design of the study and review of the manuscript but not with analyzing or interpreting the results.
FOOTNOTES
- Received 14 February 2018.
- Returned for modification 2 March 2018.
- Accepted 12 May 2018.
- Accepted manuscript posted online 4 June 2018.
- Copyright © 2018 American Society for Microbiology.
