Imipenem-relebactam (I-R) is a recently developed carbapenem–beta-lactamase inhibitor combination agent that can overcome carbapenem resistance, which has now emerged in Escherichia coli, including sequence type 131 (ST131) and its fluoroquinolone-resistant H30R subclone, the leading cause of extraintestinal E. coli infections globally. To clarify the likely utility of I-R for carbapenem-resistant (CR) E. coli infections in the United States, we characterized 203 recent CR clinical E. coli isolates from across the United States (years 2002 to 2017) for phylogroup, clonal group (including ST131, H30R, and the CTX-M-15-associated H30Rx subset within H30R), relevant beta-lactamase genes, and broth microdilution MICs for I-R and 11 comparator agents.
KEYWORDS: Escherichia coli, ST131, antimicrobial susceptibility, carbapenem resistance, clonality, coresistance, drug resistance mechanisms, imipenem-relebactam, molecular epidemiology, phylogenetic analysis
ABSTRACT
Imipenem-relebactam (I-R) is a recently developed carbapenem–beta-lactamase inhibitor combination agent that can overcome carbapenem resistance, which has now emerged in Escherichia coli, including sequence type 131 (ST131) and its fluoroquinolone-resistant H30R subclone, the leading cause of extraintestinal E. coli infections globally. To clarify the likely utility of I-R for carbapenem-resistant (CR) E. coli infections in the United States, we characterized 203 recent CR clinical E. coli isolates from across the United States (years 2002 to 2017) for phylogroup, clonal group (including ST131, H30R, and the CTX-M-15-associated H30Rx subset within H30R), relevant beta-lactamase genes, and broth microdilution MICs for I-R and 11 comparator agents. Overall, I-R was highly active (89% susceptible), more so than all comparators except tigecycline and colistin (both 99% susceptible). I-R’s activity varied significantly in relation to phylogroup, clonal background, resistance genotype, and region. It was greatest among phylogroup B2, ST131-H30R, H30Rx, Klebsiella pneumoniae carbapenemase (KPC)-positive, and northeast U.S. isolates and lowest among phylogroup C, New Delhi metallo-β-lactamase (NDM)-positive, and southeast U.S. isolates. Relebactam improved imipenem’s activity against CR isolates within each phylogroup—especially groups A, B1, and B2—and particularly against isolates containing KPC. I-R remained substantially active against isolates coresistant to comparator agents, albeit somewhat less so than against the corresponding susceptible isolates. These findings suggest that I-R should be useful for treating most CR E. coli infections in the United States, largely independent of coresistance, although this likely will vary in relation to the local prevalence of specific E. coli lineages and carbapenem resistance mechanisms.
INTRODUCTION
Carbapenem resistance is emerging in Escherichia coli, including its leading pandemic, multidrug-resistant (MDR) lineage, the H30R subset within sequence type 131 (ST131), i.e., ST131-H30R (1–5). Such emerging resistance threatens the utility of carbapenems as reliable fallback agents for infections due to organisms resistant to narrower-spectrum alternatives, such as extended-spectrum cephalosporins and fluoroquinolones, which in many locales are now unreliable for empirical therapy, especially in patients who are critically ill or at risk for MDR organisms (6).
Imipenem-relebactam (I-R), a recently developed carbapenem–beta-lactamase inhibitor combination agent, is active against many carbapenem-resistant (CR) E. coli strains (7–9) but has been studied minimally within ST131 and not specifically within ST131’s resistance-associated H30R1 and H30Rx subclones. Antimicrobial resistance in E. coli is highly clonal, frequently differing by ST and sometimes even by subclone within a given ST (10–12). Such lineage-specific susceptibility differences can include differences in MIC or minimum bactericidal concentration (MBC) distribution among isolates classified as resistant to a given agent, e.g., the higher fluoroquinolone MIC and MBC values observed among ST131-H30R isolates compared with those of other fluoroquinolone-resistant E. coli isolates (13, 14). Whether this phenomenon also occurs with carbapenems and I-R is unknown.
Accordingly, here, we tested I-R and comparator agents for activity against CR E. coli from across the United States and Minnesota in relation to phylogenetic and clonal background (including ST131 subclones), resistance genes, coresistance profiles, and geographical region. Specifically, we sought to determine how I-R’s activity compares with that of other agents and to what extent it varies in relation to specific bacterial characteristics.
RESULTS
Overall percent susceptible and MICs.
Among the 203 total CR E. coli isolates, the percent susceptible was higher for I-R (89%) than for any nonpotentiated carbapenem (meropenem [MEM], 75%; imipenem [IPM], 44%; ertapenem [ETP], 3%) or any other comparator except tigecycline (TGC; 99%) and colistin (CL; 99%) (Table 1). The MIC50 was 8-fold lower for (carbapenemase inhibitor containing) I-R (0.25 mg/liter) than for IPM (2.0 mg/liter).
TABLE 1.
Overall percent susceptible and MICmin, MIC50, MIC90, and MICmax for imipenem-relebactam and comparatorsa
| Agent | No. susceptible (% of 203) | MICmin | MIC50 | MIC90 | MICmax |
|---|---|---|---|---|---|
| I-R | 180 (89) | ≤0.03 | 0.25 | >256 | >256 |
| IPM | 90 (44) | 0.125 | 2 | >256 | >256 |
| MEM | 153 (75) | ≤0.03 | 0.5 | >4 | >4 |
| ETP | 7 (3) | ≤0.016 | >2 | >2 | >2 |
| GEN | 132 (65) | 0.25 | 1 | >16 | >16 |
| TZP | 56 (28) | ≤1 | 128 | >128 | >128 |
| AMK | 163 (80) | ≤0.5 | 4 | >64 | >64 |
| LVX | 40 (20) | ≤0.06 | >8 | >8 | >8 |
| TGC | 200 (99) | 0.25 | 0.5 | 8 | 8 |
| CAZ | 19 (9) | ≤0.125 | 32 | >256 | >16 |
| CL | 201 (99) | ≤0.06 | 0.125 | 8 | 8 |
| MIN | 141 (69) | 0.5 | 4 | >16 | >16 |
MICmin, lowest detected MIC; MICmax, highest detected MIC; AMK, amikacin; CAZ, ceftazidime; CL, colistin; ETP, ertapenem; GEN, gentamicin; IPM, imipenem; LVX, levofloxacin; MEM, meropenem; MIN, minocycline; TGC, tigecycline; TZP, piperacillin-tazobactam.
Phylogroup.
Percent susceptible to I-R varied significantly by phylogroup, being highest (>90% per phylogroup) in groups B1, B2, D, E, and F and lowest in groups A (73%) and C (27%) (Table 2). Relebactam’s susceptibility-restoring effect was phylogenetically distributed, being least evident among phylogroup C isolates, of which only a small fraction of IPM-resistant isolates were susceptible to I-R compared with other phylogroups, within which most (groups A, B2, D, and F) or all (groups B1 and E) IPM-resistant isolates were I-R-susceptible (Table 2).
TABLE 2.
Percent susceptible for imipenem-relebactam and imipenem by phylogroup
| Phylogroup | No. (% of 203) | No. susceptible (% of 203) |
P valuea | |
|---|---|---|---|---|
| Imipenem-relebactam | Imipenem | |||
| A | 33 (16) | 24 (73) | 7 (21) | <0.001 |
| B1 | 13 (6) | 13 (100) | 5 (39) | 0.008 |
| B2 | 80 (39) | 78 (98) | 40 (50) | <0.001 |
| C | 11 (5) | 3 (27) | 2 (18) | 1.0 |
| D | 40 (20) | 37 (93) | 22 (55) | <0.001 |
| E | 2 (1.0) | 2 (100) | 0 (0) | 0.50 |
| F | 24 (12) | 23 (96) | 14 (58) | 0.004 |
P values determined by McNemar’s test (two-tailed).
Clonal group.
Percent susceptible to I-R also varied significantly by clonal group (Table 3.). For the three STs that were represented sufficiently to support statistical analysis, percent susceptible was greatest among ST131 isolates (99%; versus all other isolates combined, P ≤ 0.01) and lowest among ST405 isolates (80%; versus all other isolates combined, P < 0.05). This pattern contrasted with the patterns observed for IPM, amikacin (AMK), levofloxacin (LVX), and CL (Table 3). Within ST131, the H30R1 and H30Rx subclones exhibited a similarly high percentage susceptible to I-R (100% and 98%, respectively), despite significant differences for susceptibility to IPM and piperacillin-tazobactam (TZP) (H30Rx higher) and AMK (H30R1 higher). Relebactam’s IPM susceptibility-restoring effect was evident within each analyzed clonal group but was greatest for the H30R1 isolates (64% increment in percent susceptible), the combined ST131 isolates (46% increment), and the combined non-H30 isolates (44% increment), whereas it was least for ST405 isolates (7% increment).
TABLE 3.
Percent susceptible for imipenem-relebactam and comparators by clonal group
| Agentb | Total (n = 203) | Sequence typed
|
ST131 subclonea
status |
Three-group P valuec | ||||
|---|---|---|---|---|---|---|---|---|
| ST131 (n = 68) | ST405 (n = 15) | ST648 (n = 15) | non-H30a (n = 141) | H30R1 (n = 22) | H30Rx (n = 40) | |||
| I-R | 180 (89) | 67 (99)** | 12 (80) | 14 (93) | 119 (84) | 22 (100) | 39 (98) | * |
| IPM | 90 (44) | 36 (53) | 11 (73)* | 11 (73)* | 56 (40) | 8 (36) | 26 (65) | * |
| MEM | 153 (75) | 51 (75) | 12 (80) | 14 (93) | 106 (75) | 15 (68) | 32 (80) | |
| ETP | 7 (3) | 1 (1.5) | 0 (0) | 0 (0) | 6 (4) | 0 (0) | 1 (2.5) | |
| GEN | 132 (65) | 47 (69) | 7 (47) | 10 (67) | 90 (64) | 13 (59) | 29 (73) | |
| TZP | 56 (28) | 22 (33) | 5 (33) | 5 (33) | 34 (24) | 4 (18) | 18 (45) | * |
| AMK | 163 (80) | 47 (69)** | 13 (87) | 12 (80) | 121 (86) | 20 (91) | 22 (55) | *** |
| LVX | 40 (20) | 4 (6)*** | 1 (7) | 3 (20) | 40 (28) | 0 (0) | 0 (0) | *** |
| TGC | 200 (99) | 68 (100) | 14 (93) | 15 (100) | 138 (98) | 22 (100) | 40 (100) | |
| CAZ | 19 (9) | 8 (12) | 2 (13) | 0 (0) | 12 (9) | 5 (23) | 2 (5) | |
| CL | 201 (99) | 67 (99) | 15 (100) | 15 (100) | 139 (99) | 22 (100) | 40 (100) | |
| MIN | 141 (69) | 58 (85)** | 5 (33)** | 10 (67) | 88 (62) | 20 (91) | 33 (83) | ** |
Non-H30, not a member of the ST131-H30R1 or H30Rx subclones (includes some ST131 strains); H30R1, fluoroquinolone resistance-associated subclone within ST131 (excludes H30Rx); H30Rx, extended-spectrum β-lactamase (ESBL) associated.
AMK, amikacin; CAZ, ceftazidime; CL, colistin; ETP, ertapenem; GEN, gentamicin; IPM, imipenem; I-R, imipenem-relebactam; LVX, levofloxacin; MEM, meropenem; MIN, minocycline; TGC, tigecycline; TZP, piperacillin-tazobactam.
P value (from chi-square test) for three-group comparisons across ST131 subclone categories. *, P < 0.05; **, P < 0.01; ***, P ≤ 0.001.
P value (from chi-square test) for indicated ST versus all other isolates combined. *, P < 0.05; **, P < 0.01; ***, P ≤ 0.001.
Resistance genotype.
Percent susceptible to I-R also varied significantly by resistance genotype, with the greatest effects (in opposite directions) observed with Klebsiella pneumoniae carbapenemase (KPC) (100% susceptible [if positive] versus 86% [if negative; not shown], P = 0.03) and New Delhi metallo-β-lactamase (NDM) (6% susceptible [if positive] versus 94% [if negative; not shown], P < 0.001) (Table 4). By contrast, isolates with OXA-48 were only moderately less likely to be I-R susceptible than others (69% versus 90%, P > 0.05), whereas those with CTX-M or CMY-2 resembled the population overall.
TABLE 4.
Percent susceptible for imipenem-relebactam and comparator agents by resistance genotype
| Agenta | Total no. (% of 203) | No. KPC (%) (n = 35) | P valueb | No. NDM (%) (n = 16) | P valueb | No. OXA-48 (%) (n = 9) | P valueb | No. CTX-M (%) (n = 84) | P valueb | No. CMY-2 (%) (n = 58) | P valueb |
|---|---|---|---|---|---|---|---|---|---|---|---|
| I-R | 180 (89) | 35 (100) | * | 1 (6) | *** | 6 (67) | * | 72 (86) | 50 (86) | ||
| IPM | 90 (44) | 0 (0) | *** | 0 (0) | *** | 1 (11) | * | 51 (60) | *** | 25 (43) | |
| MEM | 153 (75) | 19 (54) | ** | 1 (6) | *** | 7 (78) | 64 (76) | 49 (84) | |||
| ETP | 7 (3) | 0 (0) | 1 (6) | 0 (0) | 3 (4) | 4 (7) | |||||
| GEN | 132 (65) | 17 (49) | * | 4 (25) | *** | 6 (67) | 45 (54) | ** | 47 (81) | ** | |
| TZP | 56 (28) | 2 (6) | ** | 1 (6) | * | 0 (0) | 31 (37) | * | 13 (22) | ||
| AMK | 163 (80) | 27 (77) | 10 (63) | 7 (78) | 59 (70) | ** | 56 (97) | *** | |||
| LVX | 40 (20) | 10 (29) | 0 (0)* | 1 (11) | 1 (1.2) | *** | 23 (40) | *** | |||
| TGC | 200 (99) | 35 (100) | 15 (94) | 9 (100) | 84 (100) | 57 (98) | |||||
| CAZ | 19 (9) | 1 (3) | 1 (6) | 2 (22) | 2 (2) | ** | 0 (0) | ** | |||
| CL | 201 (99) | 33 (94) | ** | 16 (100) | 9 (100) | 84 (100) | 58 (100) | ||||
| MIN | 141 (69) | 28 (80) | 6 (37) | ** | 5 (56) | 51 (60) | * | 38 (65) |
AMK, amikacin; CAZ, ceftazidime; CL, colistin; ETP, ertapenem; GEN, gentamicin; IPM, imipenem; I-R, imipenem-relebactam; LVX, levofloxacin; MEM, meropenem; MIN, minocycline; TGC, tigecycline; TZP, piperacillin-tazobactam.
P value (by chi-square test) for indicated group versus all others. *, P < 0.05; **, P < 0.01; ***, P ≤ 0.001.
Geographical region.
Susceptibility varied by region for I-R and several comparators in patterns that differed between agents (see Table S1 in the supplemental material). For I-R, percent susceptible was highest among isolates from the Northeast and Central United States and lowest among those from the Southeast and South. By contrast, for MEM, TZP, and ceftazidime (CAZ), it was highest among isolates from the Southeast, and for IPM, MEM, gentamicin (GEN), TZP, and CAZ, it was low or lowest among isolates from the Northeast.
Resistance genotype also varied by region in patterns that differed by resistance gene (see Table S2 in the supplemental material). Specifically, KPC was most prevalent in the Northeast and least prevalent in the central region, CTX-M was most prevalent in the West (albeit with small numbers) and least prevalent in the Northeast and Southeast, and CMY-2 was largely confined to the central region.
Coresistance.
For each comparator agent except colistin, I-R MICs were higher (either significantly or numerically) among isolates that exhibited nonsusceptibility to the particular comparator agent (Table 5). Likewise, for all comparators except CAZ, CL, and ETP, the percent susceptible to I-R followed a similar pattern, being significantly or numerically lower among isolates that exhibited nonsusceptibility to the particular comparator agent (Table 6). Nonetheless, the percent of coresistant isolates that remained susceptible to I-R was >75% for all comparators except MEM (32/50, 64%; P ≤ 0.001) and TGC (1/3, 33%; P = 0.03).
TABLE 5.
Imipenem-relebactam MICs among Escherichia coli isolates susceptible or nonsusceptible to comparator agents
| Comparator agenta | I-R MIC |
P valueb | |||||||
|---|---|---|---|---|---|---|---|---|---|
| Susceptible to comparator agent |
Nonsusceptible to comparator agent |
||||||||
| MIC50 | MIC90 | Range | Mean | MIC50 | MIC90 | Range | Mean | ||
| IPM | 0.25 | 0.5 | ≤0.03–1.0 | 0.27 | 0.25 | 32 | 0.06 to >256 | 19.3 | *** |
| MEM | 0.25 | 1.0 | ≤0.03–2 | 0.35 | 0.5 | 64 | 0.12 to >256 | 43.1 | *** |
| ETP | 1.0 | 2 | 0.12–32 | 5.2 | 0.25 | 2 | ≤0.03 to >256 | 11.1 | |
| GEN | 0.25 | 1.0 | ≤0.03 to >256 | 9.1 | 0.25 | 32 | 0.06 to >256 | 14.1 | |
| TZP | 0.25 | 1.0 | 0.06–2 | 0.32 | 0.25 | 32 | ≤0.03 to >256 | 15.0 | * |
| AMK | 0.25 | 1 | ≤0.03 to >256 | 9.0 | 0.5 | 32 | 0.06 to >256 | 19.0 | ** |
| LVX | 0.25 | 1 | 0.06–4 | 0.41 | 0.25 | 8 | ≤0.03 to >256 | 14.0 | |
| TGC | 0.25 | 2 | ≤0.03 to >256 | 8.5 | 2 | > 256 | 0.5 to >256 | 172 | * |
| CAZ | 0.5 | 2 | 0.125–2 | 0.59 | 0.25 | 4 | 0.25 to >256 | 12.0 | |
| CL | 0.25 | 2 | ≤0.03 to >256 | 11.0 | 0.5 | 0.50 | 0.125–0.5 | 0.31 | |
| MIN | 0.25 | 1.0 | 0.06–64 | 2.0 | 0.25 | 64 | ≤0.03 to >256 | 31.0 | * |
AMK, amikacin; CAZ, ceftazidime; CL, colistin; ETP, ertapenem; GEN, gentamicin; IPM, imipenem; I-R, imipenem-relebactam; LVX, levofloxacin; MEM, meropenem; MIN, minocycline; TGC, tigecycline; TZP, piperacillin-tazobactam.
P values as determined by Mann-Whitney test. *, P < 0.05; **, P < 0.01; ***, P ≤ 0.001.
TABLE 6.
Percent susceptible to imipenem-relebactam among Escherichia coli isolates susceptible or nonsusceptible to comparator agents
| Comparator agenta | I-R susceptible, proportion (%) |
P valueb | |
|---|---|---|---|
| If susceptible to comparator agent | If nonsusceptible to comparator agent | ||
| IPM | 90/90 (100) | 90/113 (80) | *** |
| MEM | 148/153 (97) | 32/50 (64) | *** |
| ETP | 5/7 (71) | 175/196 (89) | |
| GEN | 124/132 (94) | 56/71 (79) | ** |
| TZP | 55/56 (98) | 125/147 (85) | ** |
| AMK | 149/163 (91) | 31/40 (78) | * |
| LVX | 39/40 (98) | 141/163 (87) | * |
| TGC | 179/200 (90) | 1/3 (33) | * |
| CAZ | 17/19 (89) | 163/184 (89) | |
| CL | 178/201 (89) | 2/2 (100) | |
| MIN | 133/141 (94) | 47/62 (76) | *** |
AMK, amikacin; CAZ, ceftazidime; CL, colistin; ETP, ertapenem; GEN, gentamicin; IPM, imipenem; I-R, imipenem-relebactam; LVX, levofloxacin; MEM, meropenem; MIN, minocycline; TGC, tigecycline; TZP, piperacillin-tazobactam.
P values as determined by chi-square test. *, P < 0.05; **, P < 0.01; ***, P ≤ 0.001.
DISCUSSION
Our study of the in vitro activity of I-R and relevant comparators against 203 CR clinical E. coli isolates from the United States yielded four main findings, which have potentially important basic and practical implications. First, I-R was highly active overall and exhibited a higher percent susceptible than all comparators except TGC and CL. Second, I-R’s activity varied significantly in relation to phylogroup, clonal background, resistance genotype, and region, being greatest among phylogroup B2, ST131-H30R, H30Rx, KPC-positive, and northeast U.S. isolates and lowest among phylogroup C, NDM-positive, and southeast U.S. isolates. Third, relebactam improved the activity of IPM against CR isolates within each phylogroup—especially groups A, B1, and B2—and particularly those containing KPC. Fourth, I-R remained substantially active against isolates coresistant to comparator agents, albeit somewhat less so than against the corresponding susceptible isolates. These findings provide guidance regarding the potential utility of I-R for both empirical and definitive therapy (15), which likely will become increasingly relevant as the prevalence of carbapenem resistance predictably rises within E. coli (5).
The present study population is unique. It represents, to our knowledge, the largest and best characterized collection of CR E. coli isolates reported to date. Previous studies have demonstrated broad activity for I-R against MDR E. coli isolates generally (7–9). However, due to the comparative rarity thus far of carbapenem resistance in E. coli, such studies included too few CR E. coli isolates for meaningful analysis of relevant variables, such as phylogenetic and clonal background, coresistance, and geographical origin. Thus, the present study fills a critical knowledge gap.
Our first main finding, i.e., the overall broad activity of I-R, provides reassurance that I-R should be useful generally in confronting CR E. coli. Indeed, the only comparator agents that demonstrated broader or more potent activity (tigecycline and colistin) are handicapped by concerns about their clinical efficacy (both agents), poor urinary tract penetration (tigecycline), toxicity (colistin), and optimal dosing (colistin).
The second main finding, i.e., significant variation in I-R activity in relation to multiple bacterial and source variables, adds nuance to the first finding. That is, it supports that E. coli should not be regarded as a homogeneous entity but rather as representing multiple discrete subcomponents that differ for, among other properties, response to I-R (novel to this study) and other antimicrobial agents (10, 11). Thus, a “one size fits all” approach risks false or nongeneralizable conclusions, depending on the admixture of E. coli variants that might occur in a given context. Interestingly, here, the ST131-H30R subclones (H30R1 and H30Rx) proved to actually be more susceptible to I-R than other isolates, which contrasts with their reputation for being more extensively and intensively antimicrobial resistant than other E. coli isolates (12–14, 16).
The third main finding confirms that relebactam’s effect in enhancing carbapenem susceptibility, although appreciable to some degree regardless of associated bacterial characteristics and locale, is strongest among isolates with KPC, a serine beta-lactamase (relebactam’s main target) (9), and weakest among those with NDM, a metalloenzyme (against which relebactam is inactive). Geographical variation in the distribution of resistance mechanisms and clonal variants likely underlies the observed regional differences in I-R activity, i.e., greatest in the northeast and lowest in the southeast United States.
The fourth main finding, i.e., that I-R’s activity was generally maintained despite coresistance to diverse comparators, suggests that I-R likely will be a useful alternative for many extensively MDR E. coli isolates (6). The observed lower activity of I-R among isolates resistant to several non-beta-lactam agents suggests that such strains possess resistance mechanisms (e.g., efflux pumps, alternative [potentially hyperexpressed] beta-lactamases, or permeability alterations) that impair the activity of I-R. This serves as a reminder of the potential for use of any antimicrobial to coselect for reduced susceptibility or frank resistance to chemically and mechanistically unrelated agents.
Study limitations include the study’s convenience sample nature, the different sampling approaches used for the two components of the study (i.e., all presumptive CR isolates from Minnesota versus the CR subset within the “all comer” isolates submitted by participating SENTRY centers), the absence of associated epidemiological and clinical data, the focus on U.S. isolates, and the small numbers in many subgroups. Study strengths include the comparatively large overall study population; attention to phylogenetic background, resistance genes, coresistance, and geography; and extensive statistical analyses of I-R susceptibility in relation to these variables.
In conclusion, we found that I-R was broadly active in vitro against CR E. coli clinical isolates from across the United States and that its activity varied in relation to multiple bacterial variables and geographical region. Notably, the H30R1 and H30Rx ST131 subclones were especially susceptible to I-R, which contrasts with their tendency to be more intensely fluoroquinolone resistant than other fluoroquinolone-resistant E. coli isolates. These findings suggest that I-R should be useful for treating CR E. coli infections in the United States, largely independent of coresistance, although this likely will vary in relation to the local prevalence of specific E. coli lineages and carbapenem resistance mechanisms.
MATERIALS AND METHODS
Study isolates.
A total of 312 presumptive CR clinical E. coli isolates from across the United States were obtained from the Minnesota Department of Health (MDH) (n = 250; isolation years, 2009 to 2017 [median, 2015]) and JMI Laboratories (n = 62; isolation years, 2002 to 2017 [median, 2013]). They represented all CR E. coli isolates available from these two reference laboratories as of study onset.
The MDH isolates had been collected as part of statewide public health surveillance for CR Enterobacteriaceae, which included voluntary (pre-2016) or mandatory (as of 2016) reporting by all clinical laboratories in the state and submission of the corresponding isolates. The isolates had been classified as CR by the submitting laboratories according to then-current definitions, which varied by year and relied on the results of phenotypic +/− molecular tests done in the submitting clinical laboratories, which varied by laboratory. By contrast, the JMI isolates represented all E. coli isolates held by the epidemiologically representative SENTRY Antimicrobial Surveillance Program (which has contributing centers distributed across the United States [17]) that exhibited resistance to at least one carbapenem according to the standardized broth microdilution testing performed by JMI Laboratories for all SENTRY isolates.
Susceptibility testing.
In the research laboratory, all 312 presumptive CR E. coli isolates underwent species identity confirmation based on colony morphology after growth on T7 agar and, as needed, Gram stain characteristic, indole production, citrate utilization, and API 20E profile (bioMérieux). Confirmed E. coli isolates underwent broth microdilution MIC determinations with three carbapenems, including ertapenem (ETP), imipenem (IPM), and meropenem (MEM), I-R, and eight noncarbapenem comparators, including amikacin (AMK), ceftazidime (CAZ), colistin (CL), gentamicin (GEN), levofloxacin (LVX), minocycline (MIN), tigecycline (TGC), and piperacillin-tazobactam (TZP). Test methods and reference strains were per the Clinical and Laboratory Standards Institute (CLSI) (18); interpretive criteria were per CLSI and the U.S. Food and Drug Administration. This testing identified 203 U.S.-source clinical E. coli isolates that were nonsusceptible to ≥1 carbapenem (median year of isolation, 2015 [range, 2002 to 2017]), including 141 from MDH (diverse Minnesota counties) and 62 from JMI Laboratories (diverse U.S. locales); these constituted the study population. Of the 203 isolates, 94 were nonsusceptible only to ertapenem, 102 were nonsusceptible to ertapenem plus imipenem and/or meropenem, and 7 were nonsusceptible only to imipenem; none were nonsusceptible only to meropenem.
Molecular typing.
Established PCR-based assays were used to define E. coli phylogroup (A, B1, B2, C, D, E, F) (19); selected STs associated with multidrug resistance, recent emergence, and/or extraintestinal infections generally (12, 20, 21); the H30 and H30Rx clonal subset within ST131 (22); and resistance genotypes for KPC, NDM, OXA, VIM, IMP, CTX-M, and CMY-2 (23–25). Fluoroquinolone-resistant ST131-H30 isolates were classified as H30R, and H30R isolates that tested negative for H30Rx were classified as H30R1 (12, 26).
Statistical methods.
Comparisons involving dichotomous variables were tested using chi-square tests (including an “N-1” chi-square test for two-group comparisons [27]) for unpaired comparisons and McNemar’s test for paired comparisons. Comparisons involving continuous variables were tested using the Mann-Whitney test. For some analyses, the population was split into three comparison groups by H30 subclone status as follows: non-H30 (whether non-ST131 or non-H30 ST131), H30R1, and H30Rx. Throughout, the significance criterion was a P value of <0.05. Due to the study’s exploratory nature, no adjustment was made for multiple comparisons.
Supplementary Material
ACKNOWLEDGMENTS
This work was supported in part by a research grant from Merck, Inc. and by the Office of Research Development, Department of Veterans Affairs. The funders had no control over the content of the report or the decision to publish.
The opinions expressed are those of the authors and not necessarily those of the authors’ institutions or the Department of Veterans Affairs.
James R. Johnson has had research grants and/or consultancies involving Achaogen, Allergan, Crucell/Janssen, Melinta, Merck, Shionogi, Syntiron, and Tetraphase. The other authors report no financial conflicts of interest.
Footnotes
Supplemental material is available online only.
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