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Carbapenem-resistant Klebsiella pneumoniae (CR-Kp) is currently a serious global concern. Antimicrobial stewardship programs (ASPs) are one of the key strategies to overcome this resistance. However, evidence about the long-term clinical and ecological impacts of ASPs is scarce. A multidisciplinary team conducted a multifaceted intervention in a CR-Kp endemic hospital over a 6-year period. We assessed the monthly long-term impacts of ASPs on carbapenem use, incidence density (ID), and crude death rates of hospital-acquired CR-Kp infections. Other variables potentially related to CR-Kp incidence and healthcare activity indicators were monitored. Carbapenem use showed a sustained reduction over the long term, with a difference of -66.19% (95% CI -87.03 to -45.34) between the expected pre-intervention trend consumption value and that obtained six years after starting the program. The ID of CR-Kp also decreased significantly and was maintained over the long term, with a relative reduction of -88.14% (95% CI; -100.4 to -75.85) at the end of the study period. The crude death rate of CR-Kp at 14 and 28 days decreased significantly after the intervention and remained steady after six years. Infection control indicator trends remained stable. This mixed ASP contributed to reducing the high incidence of infections and mortality rates of CR-Kp, achieving a sustained ecological and clinical effect.

Purpose: To characterize the resistance mechanisms affecting the cefepime-taniborbactam combination in a collection of carbapenemase-producing Enterobacterales (CPE) and carbapenem-resistant Pseudomonas spp. (predominantly P. aeruginosa; CRPA) clinical isolates., Methods: CPE (n = 247) and CRPA (n = 170) isolates were prospectively collected from patients admitted to 8 Spanish hospitals. Susceptibility to cefepime-taniborbactam and comparators was determined by broth microdilution. Cefepime-taniborbactam was the most active agent, inhibiting 97.6% of CPE and 67.1% of CRPA (MICs ≤ 8/4 mg/L). All isolates with cefepime-taniborbactam MIC > 8/4 mg/L (5 CPE and 52 CRPA) and a subset with MIC ≤ 8/4 mg/L (23 CPE and 24 CRPA) were characterized by whole genome sequencing., Results: A reduced cefepime-taniborbactam activity was found in two KPC-ST307-Klebsiella pneumoniae isolates with altered porins [KPC-62-K. pneumoniae (OmpA, OmpR/EnvZ), KPC-150-K. pneumoniae (OmpK35, OmpK36)] and one each ST133-VIM-1-Enterobacter hormaechei with altered OmpD, OmpR, and OmpC; IMP-8-ST24-Enterobacter asburiae; and NDM-5-Escherichia coli with an YRIN-inserted PBP3 and a mutated PBP2. Among the P. aeruginosa (68/76), elevated cefepime-taniborbactam MICs were mostly associated with GES-5-ST235, OXA-2+VIM-2-ST235, and OXA-2+VIM-20-ST175 isolates also carrying mutations in PBP3, efflux pump (mexR, mexZ) and AmpC (mpl) regulators, and non-carbapenemase-ST175 isolates with AmpD-T139M and PBP3-R504C mutations. Overall, accumulation of these mutations was frequently detected among non-carbapenemase producers., Conclusions: The reduced cefepime-taniborbactam activity among the minority of isolates with elevated cefepime-taniborbactam MICs is not only due to IMP carbapenemases but also to the accumulation of multiple resistance mechanisms, including PBP and porin mutations in CPE and chromosomal mutations leading to efflux pumps up-regulation, AmpC overexpression, and PBP modifications in P. aeruginosa., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Introduction: Infections caused by carbapenem-resistant Enterobacterales (CRE) and carbapenem-resistant Pseudomonas aeruginosa , including isolates producing acquired carbapenemases, constitute a prevalent health problem worldwide. The primary objective of this study was to determine the distribution of the different carbapenemases among carbapenemase-producing Enterobacterales (CPE, specifically Escherichia coli , Klebsiella pneumoniae , Enterobacter cloacae complex, and Klebsiella aerogenes ) and carbapenemase-producing P. aeruginosa (CPPA) in Spain from January 2014 to December 2018., Methods: A national, retrospective, cross-sectional multicenter study was performed. The study included the first isolate per patient and year obtained from clinical samples and obtained for diagnosis of infection in hospitalized patients. A structured questionnaire was completed by the participating centers using the REDCap platform, and results were analyzed using IBM SPSS Statistics 29.0.0., Results: A total of 2,704 carbapenemase-producing microorganisms were included, for which the type of carbapenemase was determined in 2692 cases: 2280 CPE (84.7%) and 412 CPPA (15.3%), most often using molecular methods and immunochromatographic assays. Globally, the most frequent types of carbapenemase in Enterobacterales and P. aeruginosa were OXA-48-like, alone or in combination with other enzymes (1,523 cases, 66.8%) and VIM (365 cases, 88.6%), respectively. Among Enterobacterales, carbapenemase-producing K. pneumoniae was reported in 1821 cases (79.9%), followed by E. cloacae complex in 334 cases (14.6%). In Enterobacterales, KPC is mainly present in the South and South-East regions of Spain and OXA-48-like in the rest of the country. Regarding P. aeruginosa , VIM is widely distributed all over the country. Globally, an increasing percentage of OXA-48-like enzymes was observed from 2014 to 2017. KPC enzymes were more frequent in 2017-2018 compared to 2014-2016., Discussion: Data from this study help to understand the situation and evolution of the main species of CPE and CPPA in Spain, with practical implications for control and optimal treatment of infections caused by these multi-drug resistant organisms., Competing Interests: LM-M has been a consultant for MSD, Shionogi and Fastinov, has served as speaker for Merck, Astra-Zeneca, Astellas, and Becton Dickinson and has received research support from MSD, Shionogi, Janssen-Cilag and Pfizer. JD-R is an employee of MSD Spain. RC has participated in educational programs organized by MSD, Pfizer and Shionogi and has received research support form MSD, and Venatrox. NL-E has been a consultant for MSD, Menarini, Shionogi and Fastinov and has served as a speaker for MSD, Pfizer, Menarini, Shionogi, Biomerieux and Accelerate Diagnostics. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Gracia-Ahufinger, López-González, Vasallo, Galar, Siller, Pitart, Bloise, Torrecillas, Gijón-Cordero, Viñado, Castillo-García, Campo, Mulet, Madueño-Alonso, Chamizo-López, Arrastia-Erviti, Galán-Sánchez, Fernández-Quejo, Rodríguez-Díaz, Gutiérrez-Zufiaurre, Rodríguez-Maresca, Ortega-Lafont, Yagüe-Guirao, Chaves-Blanco, Colomina-Rodríguez, Vidal-Acuña, Portillo, Franco-Álvarez de Luna, Centelles-Serrano, Azcona-Gutiérrez, Delgado-Iribarren García Campero, Rey-Cao, Muñoz, Calvo-Montes, Zboromyrska, Grandioso, Càmara, Cantón, Larrosa-Escartín, Díaz-Regañón and Martínez-Martínez.)

Objectives: CARB-ES-19 is a comprehensive, multicenter, nationwide study integrating whole-genome sequencing (WGS) in the surveillance of carbapenemase-producing K. pneumoniae (CP-Kpn) and E. coli (CP-Eco) to determine their incidence, geographical distribution, phylogeny, and resistance mechanisms in Spain., Methods: In total, 71 hospitals, representing all 50 Spanish provinces, collected the first 10 isolates per hospital (February to May 2019); CPE isolates were first identified according to EUCAST (meropenem MIC > 0.12 mg/L with immunochromatography, colorimetric tests, carbapenem inactivation, or carbapenem hydrolysis with MALDI-TOF). Prevalence and incidence were calculated according to population denominators. Antibiotic susceptibility testing was performed using the microdilution method (EUCAST). All 403 isolates collected were sequenced for high-resolution single-nucleotide polymorphism (SNP) typing, core genome multilocus sequence typing (cgMLST), and resistome analysis., Results: In total, 377 (93.5%) CP-Kpn and 26 (6.5%) CP-Eco isolates were collected from 62 (87.3%) hospitals in 46 (92%) provinces. CP-Kpn was more prevalent in the blood (5.8%, 50/853) than in the urine (1.4%, 201/14,464). The cumulative incidence for both CP-Kpn and CP-Eco was 0.05 per 100 admitted patients. The main carbapenemase genes identified in CP-Kpn were bla OXA-48 (263/377), bla KPC-3 (62/377), bla VIM-1 (28/377), and bla NDM-1 (12/377). All isolates were susceptible to at least two antibiotics. Interregional dissemination of eight high-risk CP-Kpn clones was detected, mainly ST307/OXA-48 (16.4%), ST11/OXA-48 (16.4%), and ST512-ST258/KPC (13.8%). ST512/KPC and ST15/OXA-48 were the most frequent bacteremia-causative clones. The average number of acquired resistance genes was higher in CP-Kpn (7.9) than in CP-Eco (5.5)., Conclusion: This study serves as a first step toward WGS integration in the surveillance of carbapenemase-producing Enterobacterales in Spain. We detected important epidemiological changes, including increased CP-Kpn and CP-Eco prevalence and incidence compared to previous studies, wide interregional dissemination, and increased dissemination of high-risk clones, such as ST307/OXA-48 and ST512/KPC-3., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Cañada-García, Moure, Sola-Campoy, Delgado-Valverde, Cano, Gijón, González, Gracia-Ahufinger, Larrosa, Mulet, Pitart, Rivera, Bou, Calvo, Cantón, González-López, Martínez-Martínez, Navarro, Oliver, Palacios-Baena, Pascual, Ruiz-Carrascoso, Vila, Aracil, Pérez-Vázquez, Oteo-Iglesias and the GEMARA/GEIRAS-SEIMC/REIPI CARB-ES-19 Study Group.)

Objectives: We evaluated the association of Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-Kp) rectal colonisation with crude mortality and whether this association is independent of the risk of KPC-Kp infection., Methods: This was a prospective cohort study of patients followed-up 90 days after a study of rectal colonisation. Cox regression was used to study the variables associated with crude mortality. Sensitivity analyses for 90-day crude mortality in different subcohorts were performed., Results: A total of 1244 patients (1078 non-colonised and 166 colonised) were included. None of the non-colonised patients and 78 (47.0%) of the colonised patients developed KPC-Kp infection. The 90-day crude mortality was 18.0% (194/1078) in non-colonised patients and 41.6% (69/166) in colonised patients. Rectal colonisation was not associated with crude mortality [hazard ratio (HR) = 1.03, 95% confidence interval (CI) 0.69-1.54; P = 0.85] when the model was adjusted for severe KPC-Kp infection [INCREMENT-CPE score (ICS) > 7]. KPC-Kp infection with ICS > 7 was associated with an increased risk of all-cause mortality (HR = 2.21, 95% CI 1.35-3.63; P = 0.002). In the sensitivity analyses, KPC-Kp colonisation was not associated with mortality in any of the analysed subcohorts, including patients who did not develop KPC-Kp infection (HR = 0.93, 95% CI 0.60-1.43; P = 0.74)., Conclusion: KPC-Kp rectal colonisation was not associated with crude mortality. Mortality increased when colonised patients developed severe KPC-Kp infection (ICS > 7). Rectal colonisation was a necessary although insufficient condition to die from a KPC-Kp infection., Competing Interests: Declaration of Competing Interests JRB has served as a scientific advisor for a research project for AstraZeneca, Pfizer and InfectoPharm, as a speaker in unrestricted accredited educational activities funded by Merck and has obtained funding from the Spanish Ministry of Economy and Competitiveness, the Carlos III Health Institute (ISCIII); BGG has a contract to intensify the research activity associated with the project P18/01849 (Carlos III Health Institute); JTC has served as scientific advisor for a research/consensus project for Pfizer, as an expert in a consensus document for InfectoPharm and has received payment for lectures including service on speakers bureaus and for the development of educational presentations for Pfizer, AstraZeneca, Shionogi and Merck; AC has received honoraria for the development of educational presentations for Pfizer and Shionogi. All other authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Colonization by KPC-producing Klebsiella pneumoniae (KPC-Kp) is associated with the risk of developing KPC-Kp infection. The impact of the time elapsed since a patient becomes colonized on this risk is not well known. An observational, prospective, longitudinal cohort study of colonized patients undergoing active rectal culture screening to rule out KPC-Kp colonization (July 2012 to November 2017). Patients with a positive culture at inclusion (colonized at start of follow-up) and those with a negative culture at inclusion who became colonized within 90 days (colonized during follow-up) were included in the analysis. CART analysis was used to dichotomize variables according to their association with infection. Kaplan-Meier infection-free survival curves and the log-rank test were used for group comparisons. Logistic regression was used to identify variables associated with KPC-Kp infection. Among 1310 patients included, 166 were colonized at the end of follow-up. Forty-seven out of 118 patients colonized at start of follow-up developed infection (39.8%) versus 31 out of 48 patients colonized during follow-up (64.6%; P = 0.006). Variables associated with KPC-Kp infection in the logistic regression analysis were: colonization detection during follow-up (OR, 2.74; 95% CI, 1.07 to 7.04; P = 0.03), Giannella risk score (OR, 1.51; 95% CI, 1.32 to 1.73; P < 0.001), high-risk ward (OR, 4.77; 95% CI, 1.61 to 14.10; P = 0.005) and urological manipulation after admission (OR, 3.69; 95% CI, 1.08 to 12.60; P = 0.04). In 25 out of 31 patients (80.6%) colonized during follow-up who developed KPC-Kp infection, infection appeared within 15 days after colonization. The risk of KPC-Kp infection was higher when colonization is recently acquired during hospitalization. In this prospective study, we concluded that the timing of colonization was a factor to assess when considering empirical treatment for suspected KPC-Kp infection and prophylaxis or infection control. IMPORTANCE In this study, it was confirmed that patients who became colonized during hospitalization had a higher risk of developing KPC-Kp infection than hospitalized patients who were already colonized at the start of follow-up. Besides, the risk of infection in the group of patients who became colonized during follow-up was greater in the first weeks immediately after colonization was confirmed. Our findings support the need for designing preventive strategies for patients at the highest risk of infection development, including those admitted in high-risk hospital wards and those undergoing urological procedures.

Background: Infections caused by carbapenemase-producing Enterobacterales (CPE) are not well represented in pivotal trials with ceftazidime/avibactam. The best strategy for the treatment of these infections is unknown., Methods: We conducted a multicentre retrospective observational study of patients who received ≥48 h of ceftazidime/avibactam or best available therapy (BAT) for documented CPE infections. The primary outcome was 30 day crude mortality. Secondary outcomes were 21 day clinical response and microbiological response. A multivariate logistic regression model was used to identify factors predictive of 30 day crude mortality. A propensity score to receive treatment with ceftazidime/avibactam was used as a covariate in the analysis., Results: The cohort included 339 patients with CPE infections. Ceftazidime/avibactam treatment was used in 189 (55.8%) patients and 150 (44.2%) received BAT at a median of 2 days after diagnosis of infection. In multivariate analysis, ceftazidime/avibactam treatment was associated with survival (OR 0.41, 95% CI 0.20-0.80; P = 0.01), whereas INCREMENT-CPE scores of >7 points (OR 2.57, 95% CI 1.18-1.5.58; P = 0.01) and SOFA score (OR 1.20, 95% CI 1.08-1.34; P = 0.001) were associated with higher mortality. In patients with INCREMENT-CPE scores of >7 points, ceftazidime/avibactam treatment was associated with lower mortality compared with BAT (16/73, 21.9% versus 23/49, 46.9%; P = 0.004). Ceftazidime/avibactam was also an independent factor of 21 day clinical response (OR 2.43, 95% CI 1.16-5.12; P = 0.02) and microbiological eradication (OR 0.40, 95% CI 0.18-0.85; P = 0.02)., Conclusions: Ceftazidime/avibactam is an effective alternative for the treatment of CPE infections, especially in patients with INCREMENT-CPE scores of >7 points. A randomized controlled trial should confirm these findings., (© The Author(s) 2022. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Novel β-lactam-β-lactamase inhibitor combinations currently approved for clinical use are poorly active against metallo-β-lactamase (MBL)-producing strains. We evaluated the in vitro activity of cefepime-taniborbactam (FTB [formerly cefepime-VNRX-5133]) and comparator agents against carbapenemase-producing Enterobacterales ( n  = 247) and carbapenem-resistant Pseudomonas species ( n  = 170) clinical isolates prospectively collected from different clinical origins in patients admitted to 8 Spanish hospitals. FTB was the most active agent in both Enterobacterales (97.6% MIC FTB , ≤8/4 mg/L) and Pseudomonas (67.1% MIC FTB , ≤8/4 mg/L) populations. The MIC FTB was >8 mg/L in 6/247 (2.4%) Enterobacterales isolates (3 KPC-producing Klebsiella pneumoniae isolates, 1 VIM-producing Enterobacter cloacae isolate, 1 IMP-producing E. cloacae isolate, and 1 NDM-producing Escherichia coli isolate) and in 56/170 (32.9%) Pseudomonas isolates, 19 of them carbapenemase producers (15 producers of VIM, 2 of GES, 1 of GES+VIM, and 1 of GES+KPC). Against the Enterobacterales isolates with meropenem MICs of >2 mg/L (138/247), FTB was the most active agent against both serine-β-lactamases (107/138) and MBL producers (31/138) (97.2 and 93.5% MIC FTB , ≤8/4 mg/L, respectively), whereas the activity of comparators was reduced, particularly against the MBL producers (ceftazidime-avibactam, 94.4 and 12.9%, meropenem-vaborbactam, 85.0 and 64.5%, imipenem-relebactam, 76.6 and 9.7%, ceftolozane-tazobactam, 1.9 and 0%, and piperacillin-tazobactam, 0 and 0%, respectively). Among the meropenem-resistant Pseudomonas isolates (163/170; MIC, >2 mg/L), the activities of FTB against serine-β-lactamase (35/163) and MBL (43/163) producers were 88.6 and 65.1%, respectively, whereas the susceptibilities of comparators were as follows: ceftazidime-avibactam, 88.5 and 16.0%, meropenem-vaborbactam, 8.5 and 7.0%, imipenem-relebactam, 2.9 and 2.3%, ceftolozane-tazobactam, 0 and 2.3%, and piperacillin-tazobactam, 0 and 0%, respectively. Microbiological results suggest FTB as a potential therapeutic option in patients infected with carbapenemase-producing Enterobacterales and carbapenem-resistant Pseudomonas isolates, including MBL producers.

The objective of this study was to analyse the mechanisms of resistance to carbapenems and other extended-spectrum-β-lactams and to determine the genetic relatedness of multidrug-resistant Enterobacterales (MDR-E) causing colonization or infection in solid-organ transplantation (SOT) recipients. Prospective cohort study in kidney (n = 142), liver (n = 98) or kidney/pancreas (n = 7) transplant recipients between 2014 and 2018 in seven Spanish hospitals. We included 531 MDR-E isolates from rectal swabs obtained before transplantation and weekly for 4-6 weeks after the procedure and 10 MDR-E from clinical samples related to an infection. Overall, 46.2% Escherichia coli, 35.3% Klebsiella pneumoniae, 6.5% Enterobacter cloacae, 6.3% Citrobacter freundii and 5.7% other species were isolated. The number of patients with MDR-E colonization post-transplantation (176; 71.3%) was 2.5-fold the number of patients colonized pre-transplantation (71; 28.7%). Extended-spectrum β-lactamases (ESBLs) and carbapenemases were detected in 78.0% and 21.1% of MDR-E isolates respectively. In nine of the 247 (3.6%) transplant patients, the microorganism causing an infection was the same strain previously cultured from surveillance rectal swabs. In our study we have observed a low rate of MDR-E infection in colonized patients 4-6 weeks post-transplantation. E. coli producing bla CTX-M-G1 and K. pneumoniae harbouring bla OXA-48 alone or with bla CTX-M-G1 were the most prevalent MDR-E colonization strains in SOT recipients.

Carbapenem-resistant Gram-negative bacilli (CR-GNB) are a critical public health threat, and carbapenem use contributes to their spread. Antimicrobial stewardship programs (ASPs) have proven successful in reducing antimicrobial use. However, evidence on the impact of carbapenem resistance remains unclear. We evaluated the impact of a multifaceted ASP on carbapenem use and incidence of CR-GNB in a high-endemic hospital. An interrupted time-series analysis was conducted one year before and two years after starting the ASP to assess carbapenem consumption, CR-GNB incidence, death rates of sentinel events, and other variables potentially related to CR-GNB incidence. An intense reduction in carbapenem consumption occurred after starting the intervention and was sustained two years later (relative effect -83.51%; 95% CI -87.23 to -79.79). The incidence density of CR-GNB decreased by -0.915 cases per 1000 occupied bed days (95% CI -1.743 to -0.087). This effect was especially marked in CR- Klebsiella pneumoniae and CR- Escherichia coli , reversing the pre-intervention upward trend and leading to a relative reduction of -91.15% (95% CI -105.53 to -76.76) and -89.93% (95% CI -107.03 to -72.83), respectively, two years after starting the program. Death rates did not change. This ASP contributed to decreasing CR-GNB incidence through a sustained reduction in antibiotic use without increasing mortality rates.

Enterobacteria species are common causes of hospital-acquired infections, which are associated with high morbidity and mortality rates. Immunocompromised patients such as solid organ transplant (SOT) recipients are especially at risk because they are frequently exposed to antibiotics in the course of their treatments. In this work, we used a collection of 106 Escherichia coli , 78 Klebsiella pneumoniae , 25 Enterobacter spp., and 24 Citrobacter spp. multidrug resistant strains isolated from transplant patients (hepatic, renal or renal/pancreatic) in order to examine their ability to adhere in vitro to HT-29 human colon cells, and to determine if some adhesive characteristics are associated with prevalence and persistence of these strains. A total of 33 E. coli (31%), 21 K. pneumoniae (27%), 7 Enterobacter spp. (28%), and 5 Citrobacter spp. (21%), adhered to the colon epithelial cells. Two main adherence patterns were observed in the four species analyzed, diffuse adherence, and aggregative adherence. Under transmission electronic microscopy (TEM), most bacteria lacked visible fimbria on their surface, despite their strong adherence to epithelial cells. None of the strains studied was able to induce any cytotoxic effect on HT-29 cells although some of them strongly colonizing both cells and glass coverslips at high density. Some of the strains failed to adhere to the epithelial cells but adhered strongly to the cover-slide, which shows that microscopy studies are mandatory to elucidate the adherence of bacteria to epithelial cells in vitro , and that quantitative assays using colony forming unit (CFUs) counting need to be supplemented with pictures to determine definitively if a bacterial strain adheres or not to animal cells in vitro . We report here, for the first time, the aggregative adherence pattern of two multidrug resistant (MDR) Citrobacter freundii strains isolated from human patients; importantly, biofilm formation in Citrobacter is totally dependent on the temperature; strong biofilms were formed at room temperature (RT) but not at 37°C, which can play an important role in the colonization of hospital surfaces. In conclusion, our results show that there is a great variety of adhesion phenotypes in multidrug-resistant strains that colonize transplanted patients., (Copyright © 2020 Ramos-Vivas, Chapartegui-González, Fernández-Martínez, González-Rico, Barrett, Fortún, Escudero, Marco, Linares, Nieto, Aranzamendi, Muñoz, Valerio, Aguado, Chaves, Gracia-Ahufinger, Paez-Vega, Martínez-Martínez and Fariñas.)

Objectives: To describe the case of a patient with infection due to a KPC-producing Klebsiella pneumoniae (K. pneumoniae) isolate developing ceftazidime-avibactam resistance with restored carbapenem susceptibility during ceftazidime-avibactam therapy. To review the clinical/microbiological cure and survival rates using carbapenems in other similar case reports and case series., Patients and Methods: A patient with an intra-abdominal infection due to K. pneumoniae producing the KPC-48 variant (L169P-A172T) (resistant to ceftazidime/avibactam and susceptible to carbapenems) who was treated with imipenem-cilastatin in combination with tigecycline and gentamicin. The literature was reviewed in order to summarise the in vivo (clinical/microbiological cure and survival rate) use of carbapenems in this emerging scenario., Results: The patient was successfully treated with the indicated regimen. In other reported cases (mostly with pneumonia) all-cause mortality was 50% and clinical cure was 62.5%. Meropenem-vaborbactam has been successful used in an additional case., Conclusions: A carbapenem-based regimen of combination therapy seems to be an option for treating patients infected with K. pneumoniae resistant to ceftazidime/avibactam and susceptible to carbapenems, at least when the risk of mortality is low., (Copyright © 2019 International Society for Antimicrobial Chemotherapy. Published by Elsevier Ltd. All rights reserved.)

This revision describes in detail the different diagnostic techniques of catheter-related infection, both in terms of catheter removal and preservation. Culture techniques based on catheter withdrawal are classified depending on the detection of extraluminal and/or intraluminal colonization, and new methodologies are described. In general, the most important recommendations are: (a) do not send for culture catheter tips without suspicion of infection, (b) Maki's technique is the standard for detecting extraluminal colonization, (c) take 2 pairs of peripheral blood cultures before starting antibiotic treatment, (d) use skin and connections/connectors cultures for the conservative diagnosis due to their high negative predictive value (Gram and culture), and (e) take differential quantitative blood cultures though all catheter lumens and through a peripheral vein., (Copyright © 2018 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.)

External validation of the INCREMENT-CPE risk score (ICS) for 30-day all-cause mortality is needed. There is also scarce information about whether colistin resistance influences the prognosis of carbapenem-resistant Klebsiella pneumoniae (CRKp) bacteraemia. In this study, the ability of ICS to predict all-cause mortality in the KAPECOR cohort was calculated using the area under the receiver operating characteristic (AUROC) curve. The association of colistin resistance with mortality was studied. The ICS showed an AUROC curve of 0.77 (95% CI 0.68-0.86). A cut-off of 8 points showed 96.8% sensitivity and 50.7% specificity. Mortality of low-risk patients was not different in those treated with monotherapy versus combination therapy. However, mortality of high-risk patients treated with combination therapy (37.8%) was significantly lower than in those treated with monotherapy (68.4%) (P = 0.008). To study the prognostic significance of colistin resistance, 83 selected cases of bacteraemia due to colistin-susceptible CRKp were obtained from the INCREMENT cohort for comparison. Colistin resistance could not be shown to be associated with higher mortality in either the high-risk ICS group [adjusted odds ratio (aOR) = 1.56, 95% CI 0.69-3.33; P = 0.29] or in 37 ICS-matched pairs (aOR = 1.38, 95% CI 0.55-3.42; P = 0.49), or in a sensitivity analysis including only KPC isolates (aOR = 1.81, 95% CI 0.73-4.57; P = 0.20), but the precision of estimates was low. These results validate ICS for all-cause mortality and to optimise targeted therapy for CRKp bacteraemia. Colistin resistance was not clearly associated with increased mortality., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

This study proposes an algorithm for microbiological diagnosis of urinary tract infections based on screening by luminometry and Gram-stain, followed by identification by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Positive urine samples detected with the luminometry screening Coral UTI Screen TM system underwent Gram staining and identification of the causative organism was performed by MALDI-TOF Microflex LT mass spectrometer (Bruker Daltonics, Germany). Subsequently, the results were compared with those of conventional culture identification using WIDER MIC/id system (Francisco Soria Melguizo SA, Spain). Considering the conventional approach as the gold standard, the proposed algorithm presented both a high specificity (98.1%) and a positive likelihood ratio of 37.42. The implementation of this algorithm would allow diagnosis of urinary tract infection in less than an hour in 92.4% of positive samples. This combination of techniques would be useful particularly for patients with severe UTI, pyelonephritis or urinary sepsis.

Background: The management and indication of empiric treatment in Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-Kp)-colonized patients should be improved., Methods: A prospective cohort of 94 patients colonized by KPC-Kp was followed for 90 days to validate (i) the Giannella risk score (GRS) to predict the development of any type of KPC-Kp infection and (ii) the INCREMENT-CPE score (ICS) to predict 30-day mortality in patients with infection. Both scores were combined to recommend appropriate empiric treatment. The predictive ability of the scores was measured by calculating the area under the receiver operating characteristic (AUROC) curve., Results: The GRS showed an AUROC curve for infection due to KPC-Kp of 0.92 (95% confidence interval [CI], .87-.98). The optimal cutoff point was fixed at <7 and ≥7 (92.9% sensitivity, 84.8% specificity); infection developed in 6.3% patients in the 0-6 GRS group and in 84.8% patient in the ≥7 GRS group. According to the ICS, the severity of the infection was also significantly higher in the ≥7 GRS group. The ICS showed an AUROC of 0.78 (95% CI, .65-.91) for 30-day all-cause mortality among patients with infection. A classification and regression tree analysis confirmed the GRS cutoff point at 7, and selected ≥12 points to predict a KPC-Kp infection with a high ICS., Conclusions: Our results validate the GRS and ICS for indicating empiric therapy in KPC-Kp-colonized patients.

The increasing rates of carbapenemase-producing Enterobacteriaceae (CPE) represent an important threat to health care systems and treatment of CPE infections is a challenge. The aim of the infection-carbapenem resistance evaluation surveillance trial (iCREST) was to determinate the prevalence of CPE in urine specimens in Spain and to evaluate the in vitro activity of ceftazidime-avibactam. Urine specimens (n = 11 826) were included and activity of ceftazidime-avibactam and comparators were investigated by broth microdilution in CPE. Carbapenemases were characterised by polymerase chain reaction (PCR) and sequencing as well as by whole genome sequencing (WGS). Overall prevalence of CPE was 1.6%. OXA-48 was the most prevalent (86.8%), followed by KPC (6.9%), VIM (4.8%), NDM (1.1%) and IMP (0.6%) carbapenemases. Klebsiella pneumoniae was the most common carbapenemase producer (87.8%). An uncommon carbapenemase type (IMP-8) in Spain was identify by WGS in an Enterobacter cloacae isolate, reinforcing the utility of surveillance programmes as effectives tools to detect unexpected genes that encode antimicrobial resistance. Ceftazidime-avibactam showed 100% susceptibility in KPC and OXA-48 producers and the rates of susceptibility in CPE non-susceptible to ceftazidime or meropenem were 92.1% and 96.9%, respectively. Ceftazidime-avibactam could be considered an adequate treatment option for urinary tract infections caused by KPC and OXA-48 producers., (Copyright © 2018 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.)

Combination therapy including colistin and a carbapenem has been found to be associated with lower mortality in the treatment of bloodstream infections (BSI) due to KPC-producing Klebsiella pneumoniae when the isolates show a meropenem or imipenem MIC of <16 mg/liter. However, the optimal treatment of BSI caused by colistin- and high-level carbapenem-resistant KPC-producing K. pneumoniae is unknown. A prospective cohort study including episodes of bacteremia caused by colistin-resistant and high-level meropenem-resistant (MIC ≥ 64 mg/liter) KPC-producing K. pneumoniae diagnosed from July 2012 to February 2016 was performed. The impact of combination therapy on crude 30-day mortality was analyzed by Cox regression using a propensity score as a covariate to control for indication bias and in an inverse probability of treatment weighting (IPTW) cohort. The study sample comprised 104 patients, of which 32 (30.8%) received targeted monotherapy and 72 (69.2%) received targeted combination therapy; none of them received either colistin or a carbapenem. The 30-day crude mortality rate was 30.8% (43.8% in patients treated with monotherapy and 25% in patients receiving combination therapy). In the Cox regression analysis, 30-day mortality was independently associated with septic shock at BSI onset (hazard ratio [HR], 6.03; 95% confidence interval [CI], 1.65 to 21.9; P = 0.006) and admission to the critical care unit (HR, 2.87; 95% CI, 0.99 to 8.27; P = 0.05). Targeted combination therapy was associated with lower mortality only in patients with septic shock (HR, 0.14; 95% CI, 0.03 to 0.67; P = 0.01). These results were confirmed in the Cox regression analysis of the IPTW cohort. Combination therapy is associated with reduced mortality in patients with bacteremia due to colistin-resistant KPC-producing K. pneumoniae with high-level carbapenem resistance in patients with septic shock., (Copyright © 2017 American Society for Microbiology.)

Objectives: Invasive infections caused by KPC-producing Klebsiella pneumoniae (KPCKP) are associated with very high mortality. Because infection is usually preceded by rectal colonization, we investigated if decolonization therapy (DT) with aminoglycosides had a protective effect in selected patients., Methods: Patients with rectal colonization by colistin-resistant KPCKP who were at high risk of developing infection (because of neutropenia, surgery, previous recurrent KPCKP infections or multiple comorbidities) were followed for 180 days. Cox regression analysis including a propensity score was used to investigate the impact of the use of two intestinal decolonization regimens with oral aminoglycosides (gentamicin and neomycin/streptomycin) on mortality, risk of KPCKP infections and microbiological success. The study was registered with ClinicalTrials.gov (NCT02604849)., Results: The study sample comprised 77 colonized patients, of which 44 (57.1%) received DT. At 180 days of follow-up, decolonization was associated with a lower risk of mortality in multivariate analyses (HR 0.18; 95% CI 0.06-0.55) and a lower risk of KPCKP infections (HR 0.14; 95% CI 0.02-0.83) and increased microbiological success (HR 4.06; 95% CI 1.06-15.6). Specifically, gentamicin oral therapy was associated with a lower risk of crude mortality (HR 0.15; 95% CI 0.04-0.54), a lower risk of KPCKP infections (HR 0.86; 95% CI 0.008-0.94) and increased microbiological response at 180 days of follow-up (HR 5.67; 95% CI 1.33-24.1). Neomycin/streptomycin therapy was only associated with a lower risk of crude mortality (HR 0.22; 95% CI 0.06-0.9)., Conclusions: Intestinal decolonization with aminoglycosides is associated with a reduction in crude mortality and KPCKP infections at 180 days after initiating treatment., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

The present study aimed to determine whether antiviral prevention strategies against cytomegalovirus (CMV) infection used in high-risk D+R- solid organ transplanted patients can modulate the impact of CMV replication on CD8(+) T-cell differentiation. The different CD8(+) T-cell subpopulations were measured at a single point when at least one year had elapsed since transplantation. A total of 68 D+R- patients were included, of which 33 underwent pre-emptive therapy and 35 received prophylaxis. Multivariate analysis showed that CMV replication was associated with the expansion of CD28־ EMRA CD8(+) T cells in patients managed pre-emptively but not in patients under prophylaxis (21.4% vs. 3.6%). This finding is likely related to the higher frequency of CMV recurrence observed in patients under pre-emptive therapy compared to those under prophylaxis (75% vs. 14.3%; p < 0.001). In fact, multivariate analysis showed that having more than one replication episode was associated with a 17.2% increase (p = 0.001) in the percentage of CD28־ EMRA CD8(+) T cells compared to "no episode" and with a 10.9% increase with respect to "single episodes" (p = 0.025). Additionally, patients with IFNγ response to CMV (QuantiFERON-CMV Reactive) had a higher percentage of late-differentiated CD8(+) T cells than patients lacking this response. In summary, recurrent CMV replication in D+R- patients under pre-emptive therapy was associated with the expansion of CD28־ EMRA CD8(+) T cells, which might have a short-term beneficial effect related to the high functionality of this T-cell subpopulation. Nevertheless, we cannot rule out that this accumulation might have a long-term detrimental effect related to immunosenescence and inflammation., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The aim of this study was to characterise carbapenem-resistant Klebsiella pneumoniae isolates that caused an outbreak in a hospital in the south of Spain, originating from a patient transferred in 2012 from Italy. Forty-four K. pneumoniae isolates, recovered from 28 patients, were screened by PCR for extended-spectrum β-lactamase (ESBL) and carbapenemase genes and the products were further sequenced. Plasmids were transferred by electroporation and were classified using PCR-based Inc/rep typing and IncF subtyping. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) were used to determine the genetic relatedness of the isolates. All isolates yielded positive modified Hodge test results, harboured bla(SHV-11), bla(TEM-1) and bla(KPC-3) genes, showed an identical PFGE pattern, and were assigned to clone sequence type 512 (ST512). The bla(KPC-3) gene was located on a 140-kb K2:A-:B-plasmid. In conclusion, the successful K. pneumoniae ST512 clone caused a major outbreak in Spain from an imported case and is the first description of an outbreak in this country due to the KPC-3-producing K. pneumoniae ST512 clone., (Copyright © 2014 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)

The aim of this study was to characterize timing, kinetic, and magnitude of CMV-specific immune response after hematopoietic stem cell transplantation (HSCT) and its ability to predict CMV replication and clinical outcomes. Using cell surface and intracellular cytokine staining by flow cytometry, CMV-specific T-cell response was measured in blood, while CMV viral load and chimerism were determined by real-time PCR. Patients that reconstituted CMV-specific T-cell response within 6 weeks after Allo-SCT showed a more robust immune response (CD8(+) : 0.7 cells/μl vs. 0.3/μl; P-value = 0.01), less incidence of CMV replication (33% vs. 89.5%; P-value = 0.007), reduced viral loads (1.81 log copies/ml vs. 0 copies/ml; P-value = 0.04), and better overall survival (72%; CI: 0.53-0.96 vs. 42% CI: 0.24-0.71; P-value = 0.07) than patients with a delayed immune reconstitution. Viremic patients had significantly higher transplant-related mortality than nonviremic patients after 1 year (33% CI: 0.15-0.52 vs. 0% CI: 0.05-0.34; P-value = 0.01). Risk factors independently associated with viral replication were receptor pretransplant CMV-positive serostatus (P-value = 0.02) and acquiring CMV-specific T-cell response after 6 weeks post-transplantation (P-value = 0.009). In conclusion, timing of acquiring a positive CMV-specific T-cell immune response after transplantation may identify patients with different risk for viral replication and different clinical outcomes, including survival., (© 2014 Steunstichting ESOT.)

The human JC polyomavirus (JCPyV) causes the rapidly progressing demyelinating disease progressive multifocal leukoencephalopathy (PML). The disease occurs most often in individuals with AIDS but also occurs in individuals receiving immunomodulatory therapies for immune-related diseases such as multiple sclerosis. JCPyV infection of host cells requires the pentasaccharide lactoseries tetrasaccharide c (LSTc) and the serotonin receptor 5-hydroxytryptamine (5-HT) receptor 5-HT2AR. While LSTc is involved in the initial attachment of virus to cells via interactions with VP1, the mechanism by which 5-HT2AR contributes to infection is not clear. To further define the roles of serotonin receptors in infection, HEK293A cells, which are poorly permissive to JCPyV, were transfected with 14 different isoforms of serotonin receptor. Only 5-HT2 receptors were found to support infection by JCPyV. None of the other 11 isoforms of serotonin receptor supported JCPyV infection. Expression of 5-HT2 receptors did not increase binding of JCPyV to cells, but this was not unexpected, given that the cells uniformly expressed the major attachment receptor, LSTc. Infection of these cells remained sensitive to inhibition with soluble LSTc, confirming that LSTc recognition is required for JCPyV infection. Virus internalization into HEK293A cells was significantly and specifically enhanced when 5HT2 receptors were expressed. Taken together, these data confirm that the carbohydrate LSTc is the attachment receptor for JCPyV and that the type 2 serotonin receptors contribute to JCPyV infection by facilitating entry.

Objectives: To determine the aetiology, clinical features and prognosis of CAP during the first post-pandemic influenza season. We also assessed the factors associated with severe disease and tested the ability of a scoring system for identifying influenza A (H1N1)pdm09-related pneumonia., Methods: Prospective cohort study carried out at 10 tertiary hospitals of Spain. All adults hospitalised with CAP from December 01, 2010 to March 31, 2011 were analysed., Results: A total of 747 adults with CAP required hospitalisation. The aetiology was determined in 315 (42.2%) patients, in whom 154 (21.9%) were due to bacteria, 125 (16.7%) were due to viruses and 36 (4.8%) were mixed (due to viruses and bacteria). The most frequently isolated bacteria were Streptococccus pneumoniae. Among patients with viral pneumonia, the most common organism identified were influenza A (H1N1)pdm09. Independent factors associated with severe disease were impaired consciousness, septic shock, tachypnea, hyponatremia, hypoxemia, influenza B, and influenza A (H1N1)pdm09. The scoring system evaluated did not differentiate reliably between patients with influenza A (H1N1)pdm09-related pneumonia and those with other aetiologies., Conclusions: The frequency of bacterial and viral pneumonia during the first post-pandemic influenza season was similar. The main identified virus was influenza A (H1N1)pdm09, which was associated with severe disease. Although certain presenting clinical features may allow recognition of influenza A (H1N1)pdm09-related pneumonia, it is difficult to express them in a reliable scoring system., (Copyright © 2013 The British Infection Association. Published by Elsevier Ltd. All rights reserved.)

Background: Experience with high-dose ganciclovir for the management of resistant cytomegalovirus (CMV) replication in transplant patients is limited despite its adoption as an effective therapy by some consensus documents., Methods: We studied six cases of CMV replication in solid organ transplant patients whose genotypic testing showed mutations associated with different levels of resistance to ganciclovir. All were treated with high-dose intravenous ganciclovir (7.5-10 mg/kg/12 hr) or oral valganciclovir (1350-1800 mg/12 hr) corrected according to creatinine clearance. The virologic response was considered positive if the CMV plasma viral load was undetectable. Safety was evaluated by clinical assessment, including the review of vital signs and laboratory tests., Results: All patients had asymptomatic replication, except one who had digestive disease. Four patients received universal prophylaxis with valganciclovir. Two patients received preemptive therapy with valganciclovir for individual episodes of replication. Two of the six patients received steroid boluses before the episode of replication by resistant CMV. All patients responded to treatment, including those with mutations associated with a high level of ganciclovir resistance. Four patients had neutropenia (<1.5 × 10/L), but only one received treatment., Conclusions: High-dose ganciclovir/valganciclovir can be an option in the treatment of resistant CMV replication and could be considered an alternative treatment in nonsevere patients for whom the use of foscarnet should be avoided. The toxicity of this regimen does not appear to limit its use.

B cell responses to minor histocompatibility antigens (mHags) have not been extensively studied after allogeneic hematopoietic stem cell transplantation (HSCT). Glutathione S-transferase T1 (GSTT1) is a drug metabolizing enzyme encoded by a single gene that is highly expressed in liver and kidney. Anti-GSTT1 antibodies have been described in the context of antibody-mediated rejection in kidney and liver transplantation, due to a mismatch between donor and recipient. The aim of the present study was to investigate the specific immune response against GSTT1 in HSCT with production of antibodies and their influence in the development of hepatic graft-versus-host-disease (GVHD). Forty patients and their respective donors were included in the study. The median follow-up time was 35.6 months (range 0.6-76 months) and a total of 349 serum samples were tested for the presence of anti-GSTT1 antibodies by ELISA test. Statistical analysis was performed by defining the GSTT1 null donor/positive recipient as mismatch compared with the other three genetic combinations regarded as GSTT1-matched. Antibodies were found in three patients within the group of null donor/positive recipient and one within the null/null group. Development of liver GVHD, particularly its acute form, was highly associated with the GSTT1-mismatch (P=0.0178) and with the presence of post-transplant anti-GSTT1 antibodies (P=0.0076). We conclude that GSTT1 could be considered as a new mHag in hepatic GVHD. The fact that three donors were parous females and the rapid production of antibodies after HSCT suggests the existence in the graft of memory B-cells specific for the GSTT1 antigen., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Background: Quantitative detection of cytomegalovirus (CMV) DNAemia by real-time PCR is currently the primary choice for the surveillance of active CMV infection in allogeneic stem cell transplant (Allo-SCT) recipients. Nevertheless, no universally accepted standards for CMV viral load quantitation are available, this being critical when clinical studies involving various participant centers that use different assays are planned., Objective: To compare the analytical performance of two commercially-available real-time PCR assays carried out at two different centers., Study Design: Plasma samples were collected at the University Hospital Virgen del Rocío (A) and at the Hospital Clínico Universitario (B) and were exchanged for analysis. In hospital A, DNA was extracted manually and viral loads were quantitated with the Affigene CMV Trender. In hospital B, DNA extractions were performed using an automated system and viral loads were quantitated using the CMV PCR Kit manufactured for Abbott by Qiagen., Results: A total of 80 samples obtained from Allo-SCT recipients (20 samples per each of the following CMV DNA load groups: undetectable level, <500 copies/mL, 500-5000 copies/mL, and >10,000 copies/mL) were analyzed. The Affigene CMV Trender assay yielded significantly higher viral loads than the Abbott CMV real-time PCR Kit, regardless of the DNA extraction method employed., Conclusions: Automated DNA extraction systems should be thoroughly evaluated for their analytical performance. Local guidelines for the initiation of pre-emptive therapy based on commercial real-time PCR assays measurements must be established as long as universally accepted standards for quantitative analysis of CMV DNAemia are not available., (Copyright 2010 Elsevier B.V. All rights reserved.)

Introduction: The current number of Human Immunodeficiency Virus (HIV) infected people is not known in Spain as there is no national registry. This study has aimed to estimate the prevalence of HIV infection in the population treated in a hospital emergency department (ER) as an epidemic and risk of exposure indicator during healthcare activity and to assess the differences observed regarding previous estimates., Material and Methods: We conducted a cross-sectional study of all the sera received in the ER anonymously. The final size of the pools was 5 sera. HIV antibody screening was performed using the 4th generation ELFA technique and confirmation was performed by Western Blot., Results: Seven out of the 270 pools made from 1,347 sera obtained were reactive. The individualized analysis confirmed 6 sera to be positive and 1 serum to be false positive. The observed prevalence was 0.52% (95% CI 0.10-0.94). Prevalence fell 0.87% in comparison to the years 1990-1991, although this was not statistically significant (p = 0.08)., Discussion: The implementation of HIV antibodies detection through a system of pooled batches in samples collected in the ER make it possible to assess the prevalence of infection with this virus, decreasing costs with regard to individualized analysis of sera in both economic terms as well as samples handling.

Background and Objective: Diagnosis of viral gastroenteritis is an important subject in clinical virology which is mainly determined by the availability of reagents in laboratories, such as in the case of astrovirus. The aim of this study was to estimate the increase in the diagnostic performance achieved after the incorporation of astrovirus search in the diagnosis protocol of acute viral gastroenteritis. We also analyzed the trend of infections in other more commonly searched virus, such as rotavirus and enteric adenovirus., Material and Methods: Retrospective study during 20 years that included 12,980 stool samples processed for gastroenteritis virus diagnosis. Since 1997 an enzyme immunoassay for astrovirus has been applied to those samples that are negative for rotavirus and adenovirus. The study was divided in two periods (1986-1996 and 1997-2005, without and with astrovirus diagnosis) and the percentage of patients diagnosed in each period was compared. The trend of positive results as well as the percentage of positive results over all patients studied was modelled using the least squares method., Results: The percentages of positive patients for rotavirus, adenovirus and astrovirus were 10.3%, 2.3% and 6.0% respectively, and there were uncommon co-infections by rotavirus and adenovirus (0.2%). The protocol applied to the astrovirus diagnosis increased the diagnosis rate up to 16.8% of the studied cases. Significant statistical differences were observed between the 2 study periods. A quadratic growth was observed in the results of positive diagnosis of viral gastroenteritis during the study period., Conclusions: The search of astrovirus in gastroenteritis cases by a selective protocol increased the diagnostic performance of gastrointestinal virus by 6%. In view of these results, it would be useful to implement astrovirus diagnosis in faeces with liquid or semi-liquid consistency when rotavirus and adenovirus detection is negative.