Clinical characteristics and outcome of bacteraemia caused by Enterobacter cloacae and Klebsiella aerogenes: more similarities than differences

Journal of Global Antimicrobial Resistance 25 (2021) 351–358 Contents lists available at ScienceDirect Journal of Global Antimicrobial Resistance journal homepage: www.elsevier.com/locate/jgar Clinical characteristics and outcome of bacteraemia caused by Enterobacter cloacae and Klebsiella aerogenes : more similarities than differences Rocío Álvarez-Marín a , ∗, José Antonio Lepe a , Oriol Gasch-Blasi b , José Manuel Rodríguez-Martínez c , Jorge Calvo-Montes d , Rosario Lara-Contreras e , Cecilia Martín-Gandul a , Fe Tubau-Quintano f , María Eliecer Cano-García d , Fernando Rodríguez-López g , Jesús Rodríguez-Baño h , Miquel Pujol-Rojo i , Julián Torre-Cisneros e , Luis Martínez-Martínez d , j , k , Álvaro Pascual-Hernández c , Manuel E. Jiménez-Mejías a , Spanish Network for Research in Infectious Diseases/Enterobacter spp. Bacteriemia Project Group 1 a Clinical Unit of Infectious Diseases, Microbiology and Preventive Medicine, Infectious Diseases Research Group, Institute of Biomedicine of Seville (IBiS), University of Seville/CSIC/Virgen del Rocío University Hospital, Seville, Spain b Infectious Diseases Service, Hospital Universitari Parc Taulí, Institut d’Investigació i Innovació Parc Taulí (l3PT), Sabadell, Spain, Spanish Network for Research in Infectious Diseases c Department of Microbiology, Virgen Macarena University Hospital, Seville, Spain, Infectious Diseases Research Group, Institute of Biomedicine of Seville (IBiS), University of Seville/CSIC, Seville, Spain d Department of Microbiology, Marqués de Valdecilla University Hospital –IDIVAL, Santander, Spain e Maimonides Biomedical Research Institute of Cordoba (IMIBIC), Clinical Unit of Infectious Diseases, Reina Sofia University Hospital, University of Cordoba, Córdoba, Spain f Department of Microbiology, University Hospital of Bellvitge, Barcelona, Spain, CIBER of Respiratory Diseases (CIBERes), Instituto de Salud Carlos III, Madrid, Spain g Department of Microbiology, Reina Sofía University Hospital, Maimonides Biomedical Research Institute of Cordoba (IMIBIC)/University of Cordoba, Córdoba, Spain h Department of Medicine, Clinical Unit of Infectious Diseases, Microbiology and Preventive Medicine, Virgen Macarena University Hospital, Institute of Biomedicine of Seville (IBiS), University of Seville/CSIC, Seville, Spain i Department of Infectious Diseases, Hospital Universitari de Bellvitge, Institut Català de la Salut (ICS-HUB), Spanish Network for Research in Infectious Diseases (REIPI RD12/0015), Instituto de Salud Carlos III (ISCIII), Madrid, Spain, Institut d’Investigació Biomédica de Bellvitge (IDIBELL), Barcelona, Spain j Department of Molecular Biology, University of Cantabria, Santander, Spain k Department of Microbiology, Reina Sofía University Hospital, Maimonides Biomedical Research Institute of Cordoba (IMIBIC)/University of Cordoba, Córdoba, Spain a r t i c l e i n f o Article history: Received 22 December 2020 Revised 28 March 2021 Accepted 14 April 2021 Available online 6 May 2021 Editor: Dr R. Watkins Keywords: Enterobacter cloacae Klebsiella aerogenes Enterobacter aerogenes Bacteraemia Case–control study Quinolone resistance mechanism a b s t r a c t Objectives: The genus Enterobacter is a common cause of nosocomial infections. Historically, the most frequent Enterobacter species were those of Enterobacter cloacae complex and Enterobacter aerogenes. In 2019, E. aerogenes was re-classified as Klebsiella aerogenes owing to its higher genotypic similarity with the genus Klebsiella . Our objective was to characterise and compare the clinical profiles of bacteraemia caused by E. cloacae and K. aerogenes. Methods: This 3-year multicentre, prospective cohort study enrolled consecutive patients with bacteraemia by E. cloacae or K. aerogenes . Baseline characteristics, bacteraemia features (source, severity, treatment), antibiotic susceptibility, resistance mechanisms and mortality were analysed. Results: The study included 285 patients with bacteraemia [196 (68.8%) E. cloacae and 89 (31.2%) K. aerogenes ] . The groups showed no differences in age, sex, previous use of invasive devices, place of acquisition, sources or severity at onset. The Charlson score was higher among patients with E. cloacae bacteraemia [2 (1–4) vs. 1 (0.5–3); P = 0.018], and previous antibiotic therapy was more common in patients with K. ∗Corresponding author. Permanent address: Avda. Manuel Siurot s/n 41013 Seville, Spain. Tel.: + 34 955 012 185. E-mail address: [email protected] (R. Álvarez-Marín). 1 Members of the Spanish Network for Research in Infectious Diseases/ Enterobacter spp. Bacteriemia Project Group are listed in the Acknowledgments. https://doi.org/10.1016/j.jgar.2021.04.008 2213-7165/© 2021 The Authors. Published by Elsevier Ltd on behalf of International Society for Antimicrobial Chemotherapy. This is an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/ ) R. Álvarez-Marín, J.A. Lepe, O. Gasch-Blasi et al. Journal of Global Antimicrobial Resistance 25 (2021) 351–358 aerogenes bacteraemia (57.3% vs. 41.3%; P = 0.01). Mortality was 19.4% for E. cloacae and 20.2% for K. aerogenes ( P = 0.869). Antibiotic susceptibility was similar for both species, and the incidence of multidrug resistance or ESBL production was low (6% and 5.3%, respectively), with no differences between species. Conclusion: Bacteraemias caused by E. cloacae and K. aerogenes share similar patient profiles, presentation and prognosis. Patients with E. cloacae bacteraemia had more co-morbidities and those with K. aerogenes bacteraemia had received more antibiotics. © 2021 The Authors. Published by Elsevier Ltd on behalf of International Society for Antimicrobial Chemotherapy. This is an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/ ) 1. Introduction The genus Enterobacter belongs to the family Enterobacteriaceae within the order Enterobacterales [ 1 , 2 ], and some of the species it includes have emerged in recent years as important nosocomial pathogens. Enterobacter species can cause many different infections, usually associated with invasive devices or procedures [ 3 , 4 ], and they are among the main aetiologies of nosocomial bloodstream infections (BSIs) [5–7] . Additionally, Enterobacter spp. are included in the ESKAPE ( Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter spp.) group, defined as the leading aetiologies of hospital-acquired infections caused by resistant pathogens [8] . Until recent taxonomic changes, the genus Enterobacter included 22 species sharing diverse phenotypic and genotypic features such as motility facilitated by flagella or the presence of an inducible constitutive AmpC β-lactamase [9] . Among these 22 species, the most frequent in the clinical setting (90–99% of cases) are those belonging to the Enterobacter cloacae complex and Enterobacter aerogenes [ 9 , 10 ] . However, E. aerogenes has recently been reclassified as Klebsiella aerogenes [9] . This taxonomic change was based on the genotypic characteristics of this species, as fullgenome sequence analyses have revealed that the closest species was Klebsiella pneumoniae [ 9 , 11 ]. The new classification separates these species, which have usually been studied together in the clinical setting. Most clinical studies of Enterobacter infections have included both E. cloacae and E. aerogenes without looking for or describing relevant differences between the two [12–14] . Our group recently published the largest European cohort study of Enterobacter bacteraemias describing risk factors for the development of this infection and those associated with its mortality [7] . That study included patients with E. cloacae and K. aerogenes bacteraemia, but their clinical characteristics were not analysed separately. Given the new taxonomic classification, we believe that this analysis is relevant to clarify the potential impact of their genotypic differences on clinical features. The objective of the present study was to characterise and compare the risk factors, clinical features and prognosis of BSIs caused by E. cloacae and K. aerogenes. Susceptibility to antibiotics and resistance mechanisms to cephalosporins and quinolones were also studied and compared. 2. Methods 2.1. Study design and patient selection A multicentre, observational cohort study of consecutive cases of E. cloacae and K. aerogenes bacteraemia and a nested case– control study with the prospective recruitment of identified patients (post-hoc analysis [7] ) was conducted in five Spanish university hospitals with a total of 5976 beds. The local ethics committee approved the study, waiving the requirement to obtain written informed consent owing to the observational nature of the research. The study recruited all consecutive cases of E. cloacae and K. aerogenes bacteraemia in patients aged > 14 years admitted to the participating hospitals during the 3-year study period. Only the first episode of bacteraemia was included for each patient. After the inclusion of each case, the first two patients with negative blood cultures and of the same sex, age ( ±10 years) and hospitalisation area were enrolled as controls. The only exclusion criterion for the control patients was a previous episode of BSI during their ongoing hospitalisation. Patients were identified prospectively based on the daily notification of consecutive positive blood cultures by each centre’s microbiology department. Clinical information was obtained from electronic charts. Variables were recorded using a standardised form and all patients were followed-up for 30 days or until death or hospital discharge, whichever occurred first. 2.2. Variables and definitions The following data were recorded: (i) demographic characteristics (age, sex, area of hospitalisation [medical, surgical or intensive care unit (ICU)] and hospital of origin); (ii) baseline clinical conditions, including underlying chronic diseases [heart failure, chronic obstructive pulmonary disease (COPD), diabetes mellitus, liver disease, chronic renal failure, cancer, solid-organ or bone marrow transplantation, human immunodeficiency virus (HIV) infection or other causes of immunosuppression such as neutropenia], Charlson comorbidity index [15] and McCabe index [16] , previous antimicrobial therapy (in the preceding 14 days), previous chemotherapy (in the preceding 30 days), previous corticosteroid therapy (defined as ≥10 mg of prednisone or equivalent daily for > 10 days), prior admission to an oncology unit or ICU during the ongoing hospital stay, and performance of one or more invasive procedures (insertion of venous catheter, urinary catheter or nasogastric tube; mechanical ventilation; performance of gastroscopy, colonoscopy, bronchoscopy or surgical procedures) during the preceding 10 days; and (iii) clinical variables, including place of acquisition [hospital (onset > 48 h after admission or transfer from another hospital; BSIs associated with a surgical site infection if the operation took place within 30 days) [17] , healthcare-related (within 48 h from admission and: BSI associated with an ambulatory diagnostic or therapeutic invasive procedure, or outpatients carrying an indwelling urinary catheter or a venous catheter, or patients on chronic dialysis or living in a nursing home) [18] , or community (within 48 h from admission and not meeting criteria for hospital or healthcare-related acquisition)], source of the bacteraemia [18] , severity at onset of bacteraemia (Pitt bacteremia score [ 19 , 20 ]), presence of sepsis or septic shock at onset of bacteraemia [21] and Acute Physiology and Chronic Health Evaluation (APACHE) II score for ICU patients [22] , empirical or pathogen-directed treatment, appropriate empirical treatment (defined as the prescription of at least one antimicrobial active against the strain isolated in 352 R. Álvarez-Marín, J.A. Lepe, O. Gasch-Blasi et al. Journal of Global Antimicrobial Resistance 25 (2021) 351–358 Table 1 Local distribution [ n (%)] of bacteraemia cases Hospital of collection Enterobacter cloacae ( n = 196) Klebsiella aerogenes ( n = 89) P -value H. Bellvitge 66 (33.7) 24 (27.0) 0.129 H. Reina Sofía 19 (9.7) 4 (4.5) H. Virgen Macarena 21 (10.7) 18 (20.2) H. Virgen del Rocío 50 (25.5) 25 (28.1) H. Marques de Valdecilla 40 (20.4) 18 (20.2) blood cultures in the first 24 h following the onset of the BSI), length of hospital stay (from admission to death or hospital discharge), length of ICU stay (from ICU admission to ICU discharge or death) and mortality (death from any cause during the followup period). 2.3. Microbiological analysis Enterobacter cloacae and K. aerogenes strains were identified and their antimicrobial susceptibility profile was determined according to each participating centre’s clinical microbiology laboratory criteria. Isolates from each patient taken after their inclusion in the study were sent to the Virgen del Rocío University Hospital where the following analyses were performed: (i) identification by mass spectrophotometry using a microflex II system (Bruker Daltonik GmbH, Bremen, Germany), with a threshold > 2.2 for identification at species level; and (ii) antimicrobial susceptibility testing by broth microdilution based on European Committee on Antimicrobial Susceptibility Testing (EUCAST) 2011 methods and standard criteria [23] . Multidrug resistance was defined as described by Magiorakos et al. [24] . Several resistance mechanisms were studied: (i) the inductivity of AmpC was tested in all isolates using the cefoxitin–cefotaxime–imipenem antagonism test; (ii) an extended-spectrum β-lactamase (ESBL) phenotypic test was performed in strains with a minimum inhibitory concentration (MIC) of ≥0.25 mg/L, by comparing the MICs of cefotaxime, ceftazidime and cefepime alone and combined with clavulanic acid; and (iii) the genes qnrA, qnrB, qnrC, qnrD, qnrS, aac(6  )-Ib -cr, qepA and oqxAB were screened using a multiplex PCR-based technique and single PCR [25–27] . The amplicons obtained were sequenced. 2.4. Statistical analysis Categorical variables were expressed as number of cases and percentage, and continuous variables as the median and interquartile range. A bivariate analysis was performed to compare the features of E. cloacae and K. aerogenes BSIs and the features of each species with their respective controls. Categorical variables were analysed using the χ2 test or Fisher’s test, as appropriate. For comparison of continuous variables, Student’s t -test or Mann–Whitney U -test were used, as appropriate. A P- value of ≤0.05 was considered as the threshold for statistical significance. A multivariate analysis to identify risk factors for E. cloacae and K. aerogenes bacteraemia was performed by logistic regression, including only variables that were statistically significant in the univariate analysis and that did not have collinearity. Odds ratios and their respective 95% confidence intervals were calculated. All statistical analyses were performed using PASW 18.0 Statistics software (SPSS Inc., Chicago, IL, USA). 3. Results The study included 285 patients, 196 (68.8%) with E. cloacae bacteraemia and 89 (31.2%) with K. aerogenes bacteraemia. Table 1 presents the distribution of cases by site. During the 3 years of the study, a year-over-year variation was observed, which was higher in the case of E. cloacae. With regard to the distribution of species by site, the E. cloacae:K. aerogenes ratio was approximately 2.5:1 in three of the participating hospitals, whereas it was 4.75:1 at Reina Sofía Hospital and 1.16:1 at Virgen Macarena Hospital. However, overall the species distribution in the different hospitals showed no significant differences. Patient profiles were similar for most of the baseline conditions recorded, as shown in Table 2 . However, patients with E. cloacae bacteraemia had more co-morbidities: they had a higher Charlson comorbidity index and the proportion of haemodialysis patients was also higher. Prior antibiotic therapy was more common in the group of patients with K. aerogenes bacteraemia. Regarding specific families of antibiotics, previous use of carbapenems was more common among patients with K. aerogenes bacteraemia, but no differences were found for the other groups of drugs. The risk factor analysis for the acquisition of E. cloacae or K. aerogenes bacteraemia compared with their controls is provided in Tables 3 a and 3 b, respectively. An ultimately fatal or rapidly fatal category in the McCabe index, chronic haemodialysis, previous use of corticosteroids, previous ICU stay, invasive devices and previous antibiotic therapy were identified as independent risk factors for the acquisition of E. cloacae bacteraemia. For K. aerogenes , only previous ICU stay, invasive devices and previous antibiotic therapy were identified as independent risk factors for the development of bacteraemia. There were no differences in the characteristics of E. cloacae and K. aerogenes bacteraemia episodes in terms of place of acquisition, source, severity and outcome ( Table 4 ). The most common source for both groups was the use of a vascular catheter, followed by a source of unknown origin. Catheter-related E. cloacae bacteraemia was especially common in haemodialysis patients (75% of cases). There were no differences in severity or outcome between E. cloacae and K. aerogenes bacteraemia. Overall mortality was 19.6%, with no differences between patients with E. cloacae bacteraemia and those with K. aerogenes bacteraemia. Crude mortality among patients with E. cloacae infection was higher than mortality among control patients (19.4% vs . 13%; P = 0.042). Mortality did not vary significantly between patients with K. aerogenes bacteraemia and control patients (20.2% vs . 14%; P = 0.195). Susceptibility to antibiotics and resistance mechanisms are summarised in Supplementary Table S1. A higher proportion of K. aerogenes isolates were susceptible to cefepime (84.3% vs . 73.5%; P = 0.050) and imipenem (93.3% vs . 83.7%; P = 0.037). Susceptibility profiles were similar for all of the other antibiotics tested. Multidrug resistance and ESBL production were uncommon and were homogeneous in both groups. Different plasmid-mediated quinolone resistance mechanisms were detected. The resistance patterns of the isolates exhibiting these mechanisms are summarised in Supplementary Table S2. 4. Discussion The present study shows that the clinical behaviour of E. cloacae and K. aerogenes BSIs is very similar. This similarity contrasts with the microbiological differences that have led to the taxonomic reclassification of E. aerogenes to K. aerogenes. Several fea- 353 R. Álvarez-Marín, J.A. Lepe, O. Gasch-Blasi et al. Journal of Global Antimicrobial Resistance 25 (2021) 351–358 Table 2 Baseline clinical and demographic characteristics of patients with Enterobacter cloacae and Klebsiella aerogenes bacteraemia Variable E. cloacae ( n = 196) K. aerogenes ( n = 89) Bivariate analysis ( P -value) Multivariate analysis [OR (95% CI); P -value] Age (years) [median (IQR)] 67.81 (56.75–77.43) 67.22 (52.84–77.03) 0.513 – Female sex 70 (35.7) 26 (29.2) 0.282 – McCabe UF/RF 78 (39.8) 25 (28.1) 0.057 – Charlson comorbidity index [median (IQR)] 2 (1–4) 1 (0.5–3) 0.025 0.86 (0.76–0.97); 0.018 Diabetes mellitus 50 (25.5) 18 (20.2) 0.332 – Chronic hepatopathy 87 (44.4) 46 (51.7) 0.252 – COPD 26 (13.3) 12 (13.5) 0.960 – Chronic heart failure 32 (16.3) 12 (13.5) 0.538 – Chronic renal failure 108 (55.1) 44 (49.4) 0.374 – Haemodialysis 28 (14.3) 1 (1.1) < 0.001 – Solid-organ transplantation 12 (6.1) 3 (3.4) 0.405 – HIV infection 2 (1.0) 1 (1.1) 1 – Cancer 58 (29.6) 18 (20.2) 0.097 – Haematological malignancy 16 (8.2) 3 (3.4) 0.199 – Solid tumour 42 (21.4) 15 (16.9) 0.371 – Neutropenia 10 (5.1) 4 (4.5) 1 – Previous chemotherapy 20 (10.2) 5 (5.6) 0.261 – Previous corticosteroids 22 (11.2) 7 (7.9) 0.385 – Other causes of immunosuppression 6 (3.1) 1 (1.1) 0.441 – Previous conditions or procedures during the ongoing hospital stay ICU stay 72 (36.7) 40 (44.9) 0.189 – Stay in an oncology unit 17 (8.7) 6 (6.7) 0.579 – Invasive procedure 152 (77.6) 75 (84.3) 0.192 – Catheter insertion 156 (79.6) 72 (80.9) 0.798 – Urinary catheter 107 (54.6) 48 (53.9) 0.918 – Nasogastric tube 55 (28.1) 31 (34.8) 0.249 – Mechanical ventilation 52 (26.5) 33 (37.1) 0.071 – Gastroscopy 12 (6.1) 4 (4.5) 0.783 – Colonoscopy 5 (2.6) 0 (0) 0.329 – Bronchoscopy 3 (1.5) 0 (0) 0.554 – Clean surgery 29 (14.8) 15 (16.9) 0.656 – Contaminated surgery 20 (10.2) 15 (16.9) 0.113 – Dirty surgery 14 (7.1) 4 (4.5) 0.600 – Previous antibiotic therapy 81 (41.3) 51 (57.3) 0.012 1.97 (1.18–3.29); 0.010 Penicillins 45 (23.0) 27 (30.3) 0.184 – Cephalosporins 24 (12.2) 13 (14.6) 0.582 – Carbapenems 11 (5.6) 11 (12.4) 0.048 – Quinolones 8 (4.1) 3 (3.4) 1 – Other drugs 29 (14.8) 18 (20.2) 0.301 – Length of stay prior to blood culture collection (days) [median (IQR)] 7 (1–16.75) 9 (1–22) 0.104 – NOTE: Data are n (%) unless otherwise stated. OR, odds ratio; CI, confidence interval; IQR, interquartile range; UF/RF, ultimately fatal/rapidly fatal; COPD, chronic obstructive pulmonary disease; HIV, human immunodeficiency virus; ICU, intensive care unit. tures make this species phenotypically and genotypically closer to the Klebsiella genus, and various authors had proposed this taxonomic change since 1971. This species was finally reclassified in 2019 after obtaining cumulative results from full-genome sequence analysis [9] . With regard to risk factors for these infections, patients with E. cloacae bacteraemia had more co-morbidities, a higher Charlson index and a higher number were on chronic haemodialysis. On the other hand, patients with K. aerogenes bacteraemia had a higher exposure to previous antibiotic therapies, especially carbapenems. Both characteristics (co-morbidities and previous antibiotics) have been described as risk factors for the acquisition of Enterobacter spp. bacteraemia, including E. cloacae and K. aerogenes [ 28 , 29 ]. However, comparison of these cases with their controls showed that the most important risk factors for developing either E. cloacae or K. aerogenes bacteraemia were healthcare procedures (ICU stay, invasive procedures or devices, previous antibiotic therapy). Baseline conditions, classified according to the McCabe index, were a risk factor only for E. cloacae bacteraemia. This is in line with a previously published analysis of the whole cohort [7] and, along with the low proportion (4.5%) of community-acquired bacteraemia, reflects how these micro-organisms are typically associated with healthcare procedures. The clinical presentation of the BSIs caused by E. cloacae and K. aerogenes did not show any differences in terms of place of acquisition, severity, source or outcome. A difference in APACHE II score was found (only in ICU patients), with a higher median score in E. cloacae than in K. aerogenes bacteraemia patients . To our knowledge, there has only been one study comparing the clinical features of E. cloacae and K. aerogenes BSIs in which Song et al. compared 172 E. cloacae bacteraemia cases with 67 K. aerogenes bacteraemia cases (then E. aerogenes ) from a single Korean centre [10] . In their cohort, patients with E. cloacae bacteraemia also had more co-morbidities, and patients with K. aerogenes bacteraemia showed a higher incidence of septic shock, presence of invasive devices and mortality. Klebsiella aerogenes was identified as an independent risk factor for mortality in the multivariate analysis, along with septic shock and inappropriate empirical treatment. This indicated that K. aerogenes could be more virulent, although their data showed that certain baseline characteristics of patients with K. aerogenes bacteraemia may have differed from those of patients with E. cloacae bacteraemia (e.g. the presence of invasive devices was higher, potentially indicating that there was a higher proportion of patients who had previously been admitted to the ICU). In addition, because of the single-centre design of the study, the findings may have reflected the local presence of a more virulent 354 R. Álvarez-Marín, J.A. Lepe, O. Gasch-Blasi et al. Journal of Global Antimicrobial Resistance 25 (2021) 351–358 Table 3 Risk factor analysis for acquisition of Enterobacter cloacae or Klebsiella aerogenes bacteraemia compared with controls (a) Risk factors for acquisition of Enterobacter cloacae bacteraemia compared with controls Variable Patients with E. cloacae bacteraemia ( n = 196) Controls ( n = 392) Bivariate analysis ( P- value) Multivariate analysis [OR (95% CI); P- value] Age (years) [median (IQR)] 67.8 (56.7–77.4) 67.1 (55.6–76.4) 0.901 – McCabe UF/RF 78 (39.8) 118 (30.1) 0.019 1.58 (1.08–2.33); 0.019 Charlson comorbidity index [median (IQR)] 2 (1–4) 2 (1–4) 0.310 – Diabetes mellitus 50 (25.5) 104 (26.5) 0.791 – Chronic hepatopathy 87 (44.4) 173 (44.1) 0.953 – COPD 26 (13.3) 66 (16.8) 0.261 – Chronic heart failure 32 (16.3) 52 (13.3) 0.317 – Chronic renal failure 108 (55.1) 190 (48.5) 0.129 – Haemodialysis 28 (14.3) 15 (3.8) < 0.001 5.20 (2.59–10.41) < 0.001 Solid-organ transplantation 12 (6.1) 26 (6.6) 0.812 – HIV infection 2 (1.0) 2 (0.5) 0.604 – Cancer 58 (29.6) 121 (30.9) 0.751 – Haematological malignancy 16 (8.2) 32 (8.2) 1 – Solid tumour 42 (21.4) 88 (22.4) 0.779 – Neutropenia 10 (5.1) 11 (2.8) 0.157 – Previous corticosteroids 22 (11.2) 10 (2.6) < 0.001 3.81 (1.71–8.52) 0.001 Other causes of immunosuppression 6 (3.1) 6 (1.5) 0.228 – ICU stay 72 (36.7) 93 (23.7) 0.001 1.61 (1.05–2.46) 0.027 Stay in an oncology unit 17 (8.7) 26 (6.6) 0.370 – Invasive procedure 152 (77.6) 226 (57.7) < 0.001 1.71 (1.09–2.68) 0.020 Catheter insertion 156 (79.6) 222 (56.6) < 0.001 – Urinary catheter 107 (54.6) 136 (34.7) < 0.001 – Nasogastric tube 55 (28.1) 65 (16.6) 0.001 – Mechanical ventilation 52 (26.5) 49 (12.5) < 0.001 – Surgical drainage device 23 (11.7) 37 (9.4) 0.386 – Gastroscopy 12 (6.1) 17 (4.3) 0.346 – Colonoscopy 5 (2.6) 6 (1.5) 0.389 – Bronchoscopy 3 (1.5) 7 (1.8) 1 – Clean surgery 29 (14.8) 34 (8.7) 0.024 – Contaminated surgery 20 (10.2) 24 (6.1) 0.096 – Dirty surgery 14 (7.1) 19 (4.8) 0.254 – Previous antibiotic therapy 81 (41.3) 94 (24.0) < 0.001 1.77 (1.17–2.66) 0.006 Penicillins 45 (23) 48 (12.2) 0.001 – Cephalosporins 24 (12.2) 22 (5.6) 0.005 – Carbapenems 11 (5.6) 22 (5.6) 1 – Quinolones 8 (4.1) 14 (3.6) 0.759 – Other drugs 29 (14.8) 22 (5.6) < 0.001 – (b) Risk factors for acquisition of Klebsiella aerogenes bacteraemia compared with controls Variable Patients with K. aerogenes bacteraemia ( n = 89) Controls ( n = 178) Bivariate analysis ( P- value) Multivariate analysis [OR (95% CI); P -value] Age (years) [median (IQR)] 67.2 (52.8–77.0) 66.6 (55.3–75.7) 0.848 – McCabe UF/RF 25 (28.1) 57 (32.0) 0.511 – Charlson comorbidity index [median (IQR)] 1 (0.5–3) 2 (0–3) 0.392 – Diabetes mellitus 18 (20.2) 45 (25.3) 0.359 – Chronic hepatopathy 46 (51.7) 86 (48.3) 0.604 – COPD 12 (13.5) 27 (15.2) 0.713 – Chronic heart failure 12 (13.5) 17 (9.6) 0.330 – Chronic renal failure 44 (49.4) 84 (47.2) 0.729 – Haemodialysis 1 (1.1) 0 (0) 0.333 – Solid-organ transplantation 3 (3.4) 6 (3.4) 1 – HIV infection 1 (1.1) 2 (1.1) 1 – Cancer 18 (20.2) 53 (29.8) 0.096 – Haematological malignancy 3 (3.4) 9 (5.1) 0.531 – Solid tumour 15 (16.9) 44 (24.7) 0.144 – Neutropenia 4 (4.5) 3 (1.7) 0.227 – Previous corticosteroids 7 (7.9) 9 (5.1) 0.362 – Other causes of immunosuppression 1 (1.1) 4 (2.2) 0.523 – ICU stay 40 (44.9) 57 (32.0) 0.039 0.98 (0.54–1.77); 0.955 Stay in an oncology unit 6 (6.7) 10 (5.6) 0.715 – Invasive procedures 75 (84.3) 109 (61.2) < 0.001 2.45 (1.22–4.92); 0.012 Catheter insertion 72 (80.9) 109 (61.2) 0.001 – Urinary catheter 48 (53.9) 69 (38.8) 0.019 – Nasogastric tube 31 (34.8) 37 (20.8) 0.013 – Mechanical ventilation 33 (37.1) 31 (17.4) 0.001 – Surgical drainage device 7 (7.9) 16 (9.0) 0.758 – Gastroscopy 4 (4.5) 2 (1.1) 0.098 – Colonoscopy 0 (0) 2 (1.1) 0.554 – Bronchoscopy 0 (0) 4 (2.2) 0.305 – Clean surgery 15 (16.9) 22 (12.4) 0.349 – Contaminated surgery 15 (16.9) 13 (7.3) 0.016 – Dirty surgery 4 (4.5) 8 (4.5) 1 – Previous antibiotic therapy 51 (57.3) 48 (27.0) < 0.001 2.93 (1–65–5.19) < 0.001 Penicillins 27 (30.3) 22 (12.4) < 0.001 – Cephalosporins 13 (14.6) 15 (8.4) 0.120 – Carbapenems 11 (12.4) 4 (2.2) 0.001 – Quinolones 3 (3.4) 9 (5.1) 0.756 – Other drugs 18 (20.2) 11 (6.2) 0.001 – NOTE: Data are n (%) unless otherwise stated. OR, odds ratio; CI, confidence interval; IQR, interquartile range; UF/RF, ultimately fatal/rapidly fatal; COPD, chronic obstructive pulmonary disease; HIV, human immunodeficiency virus; ICU, intensive care unit. 355 R. Álvarez-Marín, J.A. Lepe, O. Gasch-Blasi et al. Journal of Global Antimicrobial Resistance 25 (2021) 351–358 Table 4 Characteristics of the presentation and outcome of bloodstream infections caused by Enterobacter cloacae and Klebsiella aerogenes Variable E. cloacae ( n = 196) K. aerogenes ( n = 89) P -value Acquisition Nosocomial 143 (73.0) 66 (74.2) 0.832 Healthcare-related 46 (23.5) 16 (18.0) 0.298 Community 7 (3.6) 7 (7.9) 0.120 Hospitalisation area Medical ward 83 (42.3) 34 (38.2) 0.519 Surgical ward 47 (24.0) 21 (23.6) 1 ICU 66 (33.7) 34 (38.2) 0.458 Source Urinary tract 26 (13.3) 18 (20.2) 0.157 Digestive tract 34 (17.3) 10 (11.2) 0.186 Respiratory tract 12 (6.1) 9 (10.1) 0.232 Catheter 61 (31.1) 26 (29.2) 0.783 Surgical site 17 (8.7) 5 (5.6) 0.476 Burn 3 (1.5) 0 (0) 0.554 Unknown 43 (21.9) 21 (23.6) 0.761 Positive blood cultures after 3 days 9 (4.6) 6 (6.7) 0.451 Positive blood cultures after 7 days 4 (2.0) 1 (1.1) 1 Polymicrobial bacteraemia 31 (15.8) 10 (11.2) 0.307 Presentation Sepsis 55 (28.1) 30 (33.7) 0.334 Septic shock 19 (9.7) 6 (6.7) 0.414 Multiorgan failure 10 (5.1) 3 (3.4) 0.761 Pitt bacteremia score [median (IQR)] 1 (0–3) 1 (0–4) 0.561 APACHE II score [median (IQR)] a 18.5 (13.7–22.2) 14 (8–18) 0.029 Mortality at 7 days 21 (10.7) 8 (9.0) 0.655 Mortality at 14 days 29 (14.8) 14 (15.7) 0.838 Mortality at 30 days 38 (19.4) 18 (20.2) 0.869 Length of hospital stay (days) [median (IQR)] 22 (10.25–41.75) 27 (13.5–49.5) 0.396 Length of ICU stay (days) [median (IQR)] a 11.5 (7–23.5) 13 (8–20) 0.982 NOTE: Data are n (%) unless otherwise stated. ICU, intensive care unit; IQR, interquartile range; APACHE, Acute Physiology and Chronic Health Evaluation. a Only among patients admitted to the ICU. clone of K. aerogenes. Our study cohort included a higher number of patients and is the largest cohort to date in which the clinical profiles of E. cloacae and K. aerogenes BSIs have been characterised and compared. Furthermore, its multicentre design should have contributed to avoid the bias derived from a single-centre design with regard to the potential presence of virulent clones of K. aerogenes . The virulence of Enterobacter spp., including K. aerogenes , remains poorly understood owing to the limited number of studies performed in this field. However, differences in the pathogenicity of K. aerogenes (formerly E. aerogenes ) and E. cloacae have been reported [30] . In particular, several authors have found some virulence-encoding genes in K. aerogenes related to bacterial adhesion and biofilm formation ( fimH, mrkD, ycfM ) [31] and genes involved in the production of siderophores ( kfu, entB, ybtS ) [32] . These genes have also been identified in K. pneumoniae [30] , but none have been described in E. cloacae or other Enterobacter species [9] . None the less, we did not find differences in the clinical course or the prognosis of K. aerogenes bacteraemia, and the multicentre design should have avoided some of the mentioned potential biases of the study by Song et al. [10] . Our findings support the view that both micro-organisms have similar virulence and clinical behaviour, but further studies are needed to clarify the role of the virulence factors of K. aerogenes and their impact on clinical features. The antibiotic susceptibility profile was similar for E. cloacae and K. aerogenes , except for a higher susceptibility of K. aerogenes to cefepime and imipenem. The incidences of multidrug resistance and ESBL-producing isolates were low, and no strains producing a carbapenemase were identified, in line with a previous cohort study that has also reported a mortality similar to ours [33] . Multidrug resistance [34] and resistance to cephalosporins [28] have been described as risk factors for mortality in patients with Enterobacter spp. (including K. aerogenes ) bacteraemia. Enterobacterales acquire resistance to quinolones through mutations in different chromosomal genes. However, several plasmidmediated resistance mechanisms have been described that do not confer clinical resistance but facilitate survival during quinolone exposure and the selection of resistant mutants [35] . The most common resistance mechanisms involve the presence of plasmidmediated quinolone resistance qnr genes (comprising five families, qnrA, qnrB, qnrC, qnrD and qnrS ), which encode proteins that protect DNA gyrase and topoisomerase IV from quinolone action. The presence of qnr genes has been described both in E. cloacae and K. aerogenes , as well as their frequent transmission together with ES- BLs [ 36 , 37 ]. Another plasmid-mediated quinolone resistance mechanism is the production of Aac(6  )-Ib-cr, which reduces the activity of fluoroquinolones through acetylation of the drug molecule [38] . In our cohort, 8/13 ESBL-producing E. cloacae isolates also showed a plasmid-mediated quinolone resistance mechanism. Notably, they all came from the same centre, suggesting horizontal transmission of strains or plasmids. As previously mentioned, not all isolates with these resistance mechanisms showed clinical resistance to ciprofloxacin. This study had several limitations. Some were inherent to its observational design and the post-hoc analysis, while others resulted from the fact that the sample size determination and variable selection were not made specifically for comparison of the clinical profiles between E. cloacae and K. aerogenes . However, to our knowledge, this is the largest study comparing these two species, and the only one with a multicentre design, which increases its strength. Additionally, the absence of a third cohort of patients with K. pneumoniae bacteraemia may in fact represent the main limitation of the study. This option was ruled out because of 356 R. Álvarez-Marín, J.A. Lepe, O. Gasch-Blasi et al. Journal of Global Antimicrobial Resistance 25 (2021) 351–358 the risk of bias irrespective of whether a retrospective cohort or a prospective cohort study design had been used. To date, there are no published clinical studies comparing these three pathogens and this is undoubtedly an area worthy of future research. The criteria established to select control patients may also represent a matter of discussion. A similar control group selection has been successfully used in previous studies of BSIs [ 7 , 39 ] but certain authors have used as controls patients with bacteraemia caused by other pathogens (e.g. Escherichia coli ) [40] . The latter option may provide a more homogeneous control group, but our strategy avoided ascribing attributable mortality of E. cloacae or K. aerogenes to other pathogens. In conclusion, BSIs caused by E. cloacae and K. aerogenes have a similar clinical presentation and prognosis. Patients with E. cloacae bacteraemia had a higher rate of co-morbidities, while those with K. aerogenes bacteraemia had a higher rate of previous antibiotic use. Risk factors for acquiring both entities were similar, namely previous antibiotic therapy and use of invasive devices in hospital settings. Acknowledgments The Spanish Network for Research in Infectious Diseases/ Enterobacter spp. Bacteremia Project Group: Rocío Álvarez- Marín [RD16/0016/0009], Jorge Calvo [RD16/0016/0007], Marina de Cueto [RD16/0016/0001], María Eliecer Cano [RD16/0016/0007], Teresa Díaz de Terán [RD16/0016/0007], Juan Domínguez- Herrera [RD16/0016/0009], Carlos Fernández Mazarrasa [RD16/0016/0007], Oriol Gasch [RD16/0016/0005], Rosario Lara- Contreras [RD16/0016/0008], José Antonio Lepe [RD16/0016/0009], Cecilia Martín-Gandul [RD16/0016/0009], Luis Martínez-Martínez [RD16/0016/0008], Dolores Navarro-Amuedo [RD16/0016/0009], Jerónimo Pachón Díaz [RD16/0016/0009], Álvaro Pascual- Hernández [RD16/0016/0001], Miquel Pujol [RD16/0016/0005], Inmaculada Puppo [RD16/0016/0009], Fernando Rodríguez- López [RD16/0016/0008], José Manuel Rodríguez-Martínez [RD16/0016/0001], Jesús Rodriguez-Baño [RD16/0016/0001], Younes Smani [RD16/0016/0009], Fe Tubau [RD16/0016/0005], Julián de la Torre-Cisneros [RD16/0016/0008], Manuel E. Jiménez- Mejías PI [RD16/0016/0009]. The authors acknowledge Graham & Chevarría SL for proofreading the English version of the paper. Funding: This work was supported by the Instituto Carlos III [grant PS09/00916], Ministerio de Ciencia e Innovación, the Plan Nacional I + D + i 2013–2016 and Instituto Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Ciencia, Innovación y Universidades, Spanish Network for Research in Infectious Diseases [REIPI, RD16/0016/0001; RD16/0016/0005; RD16/0016/0007; RD16/0016/0008; RD16/0016/0009], co-financed by the European Regional Development Fund ‘A way to achieve Europe’, Operative program Intelligent Growth 2014–2020. Competing interests: None declared. Ethical approval: The local ethics committee approved the study [Code 01/2009], waiving the need to obtain signed informed consent owing to the observational nature of the research. 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