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The widespread dissemination of carbapenem-resistant bacteria in wastewater systems, particularly from clinical sources, poses a significant public health risk. This study assessed the concentrations and distributions of extracellular DNA (exDNA) and intracellular DNA (iDNA) harboring carbapenemase genes in wastewater from six tertiary care hospitals in Germany. We collected a total of 36 samples, comprising six biological replicates from each hospital, and analysed them using quantitative real-time PCR (qPCR) and digital PCR (dPCR). The analysis targeted seven carbapenemase genes: bla NDM , bla KPC , bla IMP , bla VIM , bla OXA-23-like , bla OXA-48-like , and bla OXA-58-like across both DNA fractions. Our results revealed significant variability in the concentrations of exDNA and iDNA across the sampling sites, with iDNA typically present at higher concentrations. Using NanoDrop One spectrophotometry and the Qubit dsDNA kit, exDNA concentrations ranged from 2.7 to 7.7 ng/mL, while Qubit recorded lower values between 1.1 and 4.0 ng/mL. Conversely, iDNA concentrations were markedly higher, spanning from 42.3 to 191.7 ng/mL with NanoDrop and 12.0 to 46.5 ng/mL with Qubit, highlighting the variability between DNA types and quantification methods. Despite its lower concentrations, exDNA comprised up to 18.2 % of total DNA, highlighting its potential role in the horizontal transfer of antimicrobial resistance genes (ARGs). The study detected target ARGs in both DNA fractions at all sites, with notable differences in their concentrations; iDNA consistently exhibited higher levels of ARGs, with the highest concentrations reaching 10.57 ± 0.20 log gene copies per liter (GC/L) for bla VIM in iDNA and 6.96 ± 0.72 log GC/L for bla IMP in exDNA. dPCR demonstrated greater sensitivity than qPCR, especially effective for detecting low-abundance targets like bla OXA-23-like in the exDNA fraction. Additionally, qPCR's susceptibility to inhibition and contamination emphasizes the superior robustness of dPCR. This research highlights the need for improved monitoring and the implementation of advanced treatment technologies to mitigate ARG dissemination in wastewater., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Carbapenemase-producing Klebsiella pneumoniae (CP-Kp) isolates are a public health concern as they can cause severe hospital-acquired infections that are difficult to treat. It has recently been shown that CP-Kp can take up virulence factors from hypervirulent K. pneumoniae lineages. In this study, 109 clinical CP-Kp isolates from the University Hospital Cologne were examined for the presence of acquired virulence factors using whole-genome sequencing and phenotypic tests, and results were linked to clinical data. The virulence factor iuc was present in 18/109 of the CP-Kp isolates. Other acquired virulence factors, such as ybt , cbt , iro , rmpA/rmpA2 , peg-344 , and hypervirulence-associated capsule types were detected in various combinations among these isolates. The iuc -positive isolates produced OXA-232 ( n = 7), OXA-48 ( n = 6), OXA-48+NDM ( n = 3), NDM, and KPC (each n = 1), and 7/18 isolates were resistant to ceftazidime-avibactam, colistin, and/or cefiderocol. Four isolates carried hybrid plasmids that harbored acquired virulence factors alongside the carbapenemase genes bla NDM-1/5 or bla OXA-48 . In 15/18 patients, iuc -positive CP-Kp were isolated from a clinically manifest infection site. Among these, four patients had osteomyelitis, and four patients died from pneumonia with OXA-232-producing ST231 isolates, three of them as part of an outbreak. In conclusion, acquired virulence factors are frequently detected in various combinations in carbapenemase-producing K. pneumoniae isolates in Germany, warranting continuous monitoring of infections caused by these strains., Competing Interests: The authors declare no conflict of interest.

Objectives: Assessment of vancomycin-resistant Enterococcus faecium (VREfm) prevalence upon hospital admission and analysis of risk factors for colonization., Methods: From 2014 to 2018, patients were recruited within 72 hours of admission to seven participating German university hospitals, screened for VREfm and questioned for potential risk factors (prior multidrug-resistant organism detection, current/prior antibiotic consumption, prior hospital, rehabilitation or long-term care facility stay, international travel, animal contact and proton pump inhibitor [PPI]/antacid therapy). Genotype analysis was done using cgMLST typing. Multivariable analysis was performed., Results: In 5 years, 265 of 17,349 included patients were colonized with VREfm (a prevalence of 1.5%). Risk factors for VREfm colonization were age (adjusted OR [aOR], 1.02; 95% CI, 1.01-1.03), previous (aOR, 2.71; 95% CI, 1.87-3.92) or current (aOR, 2.91; 95% CI, 2.60-3.24) antibiotic treatment, prior multidrug-resistant organism detection (aOR, 2.83; 95% CI, 2.21-3.63), prior stay in a long-term care facility (aOR, 2.19; 95% CI, 1.62-2.97), prior stay in a hospital (aOR, 2.91; 95% CI, 2.05-4.13) and prior consumption of PPI/antacids (aOR, 1.29; 95% CI, 1.18-1.41). Overall, the VREfm admission prevalence increased by 33% each year and 2% each year of life. 250 of 265 isolates were genotyped and 141 (53.2%) of the VREfm were the emerging ST117. Multivariable analysis showed that ST117 and non-ST117 VREfm colonized patients differed with respect to admission year and prior multidrug-resistant organism detection., Discussion: Age, healthcare contacts and antibiotic and PPI/antacid consumption increase the individual risk of VREfm colonization. The VREfm admission prevalence increase in Germany is mainly driven by the emergence of ST117., (Copyright © 2022 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Introduction: Candida auris is an emerging pathogen in health care-associated infections. In contrast to many other countries with rising numbers of C. auris, only seven cases have been reported in Germany from 2015 to 2017, mostly from patients who received prior medical treatment abroad. We therefore established a mandatory screening for C. auris colonisation at our tertiary care centre for all patients who were admitted as international patients or previously hospitalised in a foreign country within the past 6 months., Methods: Colonisation of patients was assessed using a previously established screening protocol for multidrug resistant bacteria. Since 2017, all screening samples were additionally analysed for C. auris using CHROMagar Candida (CHROMagar, Paris, France). Yeast isolates were identified using matrix-assisted laser ionisation time-of-flight (MALDI TOF), except for C. albicans (identified by the typical green colour on chromogenic agar). Data were analysed retrospectively., Results: Our study cohort included 655 patients and an overall number of 1399 samples. Fifty-three patients were colonised with Candida species (C. albicans, n = 37; C. glabrata, n = 14; others n = 9). No case of C. auris was detected. Candida spp. were mainly detected from respiratory samples (5.4% positive) and gastrointestinal specimen (5.2%). Laboratory costs were 14,689 € and analyses resulted in 98.7 h of additional technician's work., Conclusion: No colonisation with C. auris was detected among patients with previous hospitalisation abroad. Universal C. auris screening of patients with any contact to foreign health care does not seem to be cost-effective in our setting and more targeted screening strategies have to be developed., (© 2022 The Authors. Mycoses published by Wiley-VCH GmbH.)

Species within the Enterobacter cloacae complex (ECC) include globally important nosocomial pathogens. A three-year study of ECC in Germany identified Enterobacter xiangfangensis as the most common species (65.5%) detected, a result replicated by examining a global pool of 3246 isolates. Antibiotic resistance profiling revealed widespread resistance and heteroresistance to the antibiotic colistin and detected the mobile colistin resistance (mcr)-9 gene in 19.2% of all isolates. We show that resistance and heteroresistance properties depend on the chromosomal arnBCADTEF gene cassette whose products catalyze transfer of L-Ara4N to lipid A. Using comparative genomics, mutational analysis, and quantitative lipid A profiling we demonstrate that intrinsic lipid A modification levels are genospecies-dependent and governed by allelic variations in phoPQ and mgrB, that encode a two-component sensor-activator system and specific inhibitor peptide. By generating phoPQ chimeras and combining them with mgrB alleles, we show that interactions at the pH-sensing interface of the sensory histidine kinase phoQ dictate arnBCADTEF expression levels. To minimize therapeutic failures, we developed an assay that accurately detects colistin resistance levels for any ECC isolate., (© 2023. The Author(s).)

To determine viral dynamics in Omicron breakthrough infections, we measured severe acute respiratory syndrome coronavirus 2 RNA in 206 double-vaccinated or boostered individuals. During the first 3 days following the onset of symptoms, viral loads were significantly higher (cycle threshold [Ct], 21.76) in vaccinated compared to boostered (Ct, 23.14) individuals (P = .029). However, by performing a longitudinal analysis on 32 individuals over 14 days, no difference in the viral load trajectory was observed between double-vaccinated and boostered patients. Our results indicate that booster immunization results in a reduction in detectable viral loads without significantly changing viral load dynamics over time., Competing Interests: Potential conflicts of interest . All authors: No reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2022. Published by Oxford University Press on behalf of Infectious Diseases Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Extreme immature infants are at an increased risk of fungal infection due to immaturity of the skin barrier and the immune system. Besides Candida infections, in particular, Aspergillus may cause life-threatening diseases in preterm infants. Frequently, Aspergillus primarily affects the skin and may cause extensive damage., Methods: We searched our hospital database for fungal infections other than Candida in preterm infants treated between 2015 and 2020 at our level III neonatal intensive care unit of the University Hospital of Cologne., Results: In total, 13 preterm infants were identified. Of these, 11 had cutaneous Aspergillosis, one infant had severe enterocolitis caused by Aspergillus and Rhizopus and one had invasive intraabdominal Trichosporon mucoides infection. All infants were born <24 weeks of gestation, were delivered due to premature labor or chorioamnionitis, and had received prenatal steroids and/or hydrocortisone. Voriconazole and liposomal Amphotericin B were first-line treatments and the length of treatment varied between 3 and 148 days. Two infants died associated with severe infection. Liver toxicity was observed in six infants treated with Voriconazole. Therapeutic drug management for Voriconazole was performed in four infants. Target levels were not achieved by the doses that are recommended., Conclusions: Rare fungal infections, predominantly cutaneous Aspergillosis affects the most immature preterm infants and may cause severe disease. Treatment with Voriconazole has a high rate of liver toxicity and target levels are difficult to achieve in extremely immature infants., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

BackgroundEvidence supporting the effectiveness of single-room contact precautions (SCP) in preventing in-hospital acquisition of vancomycin-resistant enterococci (haVRE) is limited.AimWe assessed the impact of SCP on haVRE and their transmission.MethodsWe conducted a prospective, multicentre cohort study in German haematological/oncological departments during 2016. Two sites performed SCP for VRE patients and two did not (NCP). We defined a 5% haVRE-risk difference as non-inferiority margin, screened patients for VRE, and characterised isolates by whole genome sequencing and core genome MLST (cgMLST). Potential confounders were assessed by competing risk regression analysis.ResultsWe included 1,397 patients at NCP and 1,531 patients at SCP sites. Not performing SCP was associated with a significantly higher proportion of haVRE; 12.2% (170/1,397) patients at NCP and 7.4% (113/1,531) patients at SCP sites (relative risk (RR) 1.74; 95% confidence interval (CI): 1.35-2.23). The difference (4.8%) was below the non-inferiority margin. Competing risk regression analysis indicated a stronger impact of antimicrobial exposure (subdistribution hazard ratio (SHR) 7.46; 95% CI: 4.59-12.12) and underlying disease (SHR for acute leukaemia 2.34; 95% CI: 1.46-3.75) on haVRE than NCP (SHR 1.60; 95% CI: 1.14-2.25). Based on cgMLST and patient movement data, we observed 131 patient-to-patient VRE transmissions at NCP and 85 at SCP sites (RR 1.76; 95% CI: 1.33-2.34).ConclusionsWe show a positive impact of SCP on haVRE in a high-risk population, although the observed difference was below the pre-specified non-inferiority margin. Importantly, other factors including antimicrobial exposure seem to be more influential.

Objectives: To characterize two Enterococcus faecium isolates with different resistance phenotypes obtained from the same blood culture., Methods: The isolates were identified by MALDI-TOF MS and antimicrobial susceptibility testing (AST) was performed using a VITEK® 2 AST P592 card and Etest. WGS was performed on the MiSeq and MinION sequencer platforms. Core-genome MLST (cgMLST) and seven-loci MLST were performed. Plasmid analysis was performed using S1-PFGE followed by Southern-blot hybridization., Results: Both E. faecium isolates were ST203. AST revealed that one was a vancomycin-resistant E. faecium (VREfm) isolate and the other was a vancomycin-susceptible E. faecium (VSEfm) isolate. The VREfm isolate harboured the vanA gene cluster as part of a Tn1546-type transposon encoded on a 49 kb multireplicon (rep1, rep2 and rep7a) plasmid (pAML0157.1). On the same plasmid, ant(6)-Ia, cat-like and erm(B) were encoded. The VSEfm isolate harboured a rep2 plasmid (pAML0158.1), 12 kb in size, which was present in full length as part of pAML0157.1 from the VREfm isolate. The vanA-encoding pAML0157.1 was a chimera of the rep2 pAML0158.1 and a second DNA segment harbouring vanA, ant(6)-Ia, erm(B) and cat-like, as well as the replicons rep1 and rep7a. By cgMLST analysis, the VREfm and VSEfm isolates were identical., Conclusions: Our results demonstrate that the VREfm and VSEfm blood culture isolates represented ST203 and were identical. The investigated heterogeneous resistance phenotypes resulted from the acquisition or loss of plasmid segments in the enterococcal isolates. These data illustrate that mobile genetic elements may contribute to the spread of vancomycin resistance among enterococci and to the genotypic and phenotypic variation within clonal isolates., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: Hospital-acquired infections due to vancomycin-resistant enterococci (VRE) are emerging globally. The aims of our study were to estimate VRE colonisation prevalence in patients upon admission, to determine possible risk factors for VR E. faecium acquisition that already exist in the outpatient setting, and to monitor whether VRE-colonised patients developed a VRE infection during their current hospital stay. Methods: In 2014 and 2015, patients admitted to non-intensive care units were screened for rectal VRE carriage. The study patients filled out a questionnaire on potential risk factors. Analyses were restricted to VR E. faecium carriage. All patients with VRE colonisation were retrospectively monitored for infections with VRE during their current hospital stay. Results: In 4,013 enrolled patients, the VRE colonisation prevalence upon admission was 1.2% (n=48), and colonisation prevalence was 1.1% (n=45) for VR E. faecium . Only one VRE-colonised patient developed an infection with the detection of a VRE, among others. Colonisation with VR E. faecium was associated with current antibiotic use. Risk factors of VR E. faecium colonisation upon admission were increasing age, previous colonisation or infection with multidrug resistant organisms, sampling year 2015, and, within the previous six months, antibiotic exposure, a stay at a rehabilitation center, and a hospital stay. Conclusions: We observed that antibiotic treatment which occurred prior admission influenced VR E. faecium prevalence upon admission. Thus, wise antibiotic use in outpatient settings plays a major role in the prevention of VR E. faecium acquisition., Competing Interests: The authors declare that they have no competing interests., (Copyright © 2021 Bui et al.)

Objectives: To assess the admission prevalence of third-generation cephalosporin-resistant Enterobacterales (3GCREB) and to assess whether risk factors vary by β-lactamase genotype., Methods: Adult patients were recruited within 72 h of admission to general wards of six university hospitals in 2014 and 2015. Rectal swabs were screened for 3GCREB and isolates were analysed phenotypically and genotypically. Patients were questioned on potential risk factors. Multivariable analyses were performed to identify risk factors for 3GCREB colonization and for specific β-lactamases., Results: Of 8753 patients screened, 828 were 3GCREB positive (9.5%). Eight hundred and thirteen isolates were available for genotyping. CTX-M-15 was the most common ESBL (38.0%), followed by CTX-M-1 (22.5%), CTX-M-14 (8.7%), CTX-M-27 (7.5%) and SHV-ESBL (4.4%). AmpC was found in 11.9%. Interestingly, 18 Escherichia coli isolates were AmpC positive, 12 of which (67%) contained AmpC on a gene of plasmid origin [CMY (n = 10), DHA (n = 2)]. Risk factors for 3GCREB colonization varied by genotype. Recent antibiotic exposure and prior colonization by antibiotic-resistant bacteria were risk factors for all β-lactamases except CTX-M-14 and CTX-M-27. Travel outside Europe was a risk factor for CTX-M-15 and CTX-M-27 [adjusted OR (aOR) 3.49, 95% CI 2.88-4.24 and aOR 2.73, 95% CI 1.68-4.43]. A previous stay in a long-term care facility was associated with CTX-M-14 (aOR 3.01, 95% CI 1.98-4.59). A preceding hospital stay in Germany increased the risk of CTX-M-15 (aOR 1.27, 95% CI 1.14-1.41), while a prior hospital stay in other European countries increased the risk of SHV-ESBL colonization (aOR 3.85, 95% CI 1.67-8.92)., Conclusions: The detection of different ESBL types is associated with specific risk factor sets that might represent distinct sources of colonization and ESBL-specific dissemination routes., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The coronavirus disease (COVID-19) pandemic has caused tremendous pressure on hospital infrastructures such as emergency rooms (ER) and outpatient departments. To avoid malfunctioning of critical services because of large numbers of potentially infected patients seeking consultation, we established a COVID-19 rapid response infrastructure (CRRI), which instantly restored ER functionality. The CRRI was also used for testing of hospital personnel, provided epidemiological data and was a highly effective response to increasing numbers of suspected COVID-19 cases.

Introduction. Transmission of Enterobacterales in neonatal intensive care units (NICU) can cause outbreaks of colonization and invasive infections among neonates. Two clusters of nosocomial transmission of Klebsiella pneumoniae identified by MALDI-ToF mass-spectrometry were suspected at two NICUs in July and August 2016. Aim. To assess the potential transmission of K. pneumoniae among neonates. Methodology. Whole-genome sequencing (WGS) was performed of K. pneumoniae isolates obtained through targeted surveillance of patients and environmental sampling. Results. WGS data revealed that patient and environmental isolates represented two species, K. pneumoniae and K. variicola . Core-genome multi-locus sequence typing (cgMLST) of the isolates identified three separate transmission clusters, in Hospital A a cluster of K. pneumoniae isolates in 12 children and two environmental samples and a second cluster of K. variicola isolates in five children. In Hospital B a cluster of K. pneumoniae isolates from three children and five unrelated isolates of K. pneumoniae and two unrelated isolates of K. variicola were found. Conclusion. K. variicola can cause hospital outbreaks of colonization and infection similar to other Klebsiella spp.Preliminary results from this study were presented at the 27th European Congress of Clinical Microbiology and Infectious Diseases, April 22-25, 2018, Vienna, Austria.

Objectives: Clostridioides difficile infection (CDI) is one of the most important healthcare-associated infections. We aimed to describe the incidence density of healthcare-associated CDI (HA-CDI) in Germany's largest hospital and to identify associations with ward-level antimicrobial consumption., Methods: We used surveillance data on CDI and antimicrobial consumption from 2014 to 2017 and analysed a potential association by means of multivariable regression analysis., Results: We included 77 wards with 404998 admitted patients and 1850862 patient-days. Six hundred and seventy-one HA-CDI cases were identified, resulting in a pooled mean incidence density of 0.36/1000 patient-days (IQR = 0.34-0.39). HA-CDI incidence density on ICU and haematological-oncological wards was about three times higher than on surgical wards [incidence rate ratio (IRR) = 3.00 (95% CI = 1.96-4.60) and IRR = 2.78 (95% CI = 1.88-4.11), respectively]. Ward-level consumption of third-generation cephalosporins was the sole antimicrobial risk factor for HA-CDI. With each DDD/100 patient-days administered, a ward's HA-CDI incidence density increased by 2% [IRR = 1.02 (95% CI = 1.01-1.04)]. Other risk factors were contemporaneous community-associated CDI cases [IRR = 1.32 (95% CI = 1.07-1.63)] and CDI cases in the previous month [IRR = 1.27 (95% CI = 1.07-1.51)]. Furthermore, we found a significant decrease in HA-CDI in 2017 compared with 2014 [IRR = 0.68 (95% CI = 0.54-0.86)]., Conclusions: We confirmed that ward-level antimicrobial use influences HA-CDI and specifically identified third-generation cephalosporin consumption as a risk factor., (© The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Background: Infections caused by third generation cephalosporin-resistant Enterobacteriaceae (3GCREB) are an increasing healthcare problem. We aim to describe the 3GCREB infection incidence and compare it to prevalence upon admission. In addition, we aim to describe infections caused by 3GCREB, which are also carbapenem resistant (CRE)., Methods: In 2014-2015, we performed prospective 3GCREB surveillance in clinically relevant patient specimens (screening specimens excluded). Infections counted as hospital-acquired (HAI) when the 3GCREB was detected after the third day following admission, otherwise as community-acquired infection (CAI)., Results: Of 578,420 hospitalized patients under surveillance, 3367 had a 3GCREB infection (0.58%). We observed a similar 3GCREB CAI and HAI incidence (0.28 and 0.31 per 100 patients, respectively). The most frequent pathogen was 3GCR E. coli , in CAI and HAI (0.15 and 0.12 per 100 patients). We observed a CRE CAI incidence of 0.006 and a HAI incidence of 0.008 per 100 patients (0.014 per 1000 patient days)., Conclusions: Comparing the known 3GCREB admission prevalence of the participating hospitals (9.5%) with the percentage of patients with a 3GCREB infection (0.58%), we conclude the prevalence of 3GCREB in university hospitals to be about 16 times higher than suggested when only patients with 3GCREB infections are considered. Moreover, we find the HAI and CAI incidence caused by CRE in Germany to be relatively low., Competing Interests: The ethics committee at Charité, University Medicine Berlin, Germany, approved this study (EA/018/14). Surveillance was performed in accordance with the German Infection Protection Act and did not require patient consent.(9).Not applicable.No competing interests unless stated. MJGTV is a consultant to: Alb Fils Kliniken, Astellas Pharma, MaaT Pharma, MSD/Merck; has served with the speakers’ bureau of: Astellas Pharma, Basilea, Gilead Sciences, Merck/MSD, Organobalance and Pfizer; received research funding from: 3 M, Astellas Pharma, DaVolterra, Gilead Sciences, Merck/MSD, Morphochem, Organobalance, and Seres Therapeutics. HS reports grants from Bundesministerium für Bildung und Forschung (BMBF), the German Center for Infection Research (DZIF), Cubist, and Novartis, and personal fees from Astellas-Basilea, Cubist, Durata, Genentech, Gilead, MSD, Roche Pharma, and Tetraphase. JKMK received research and travel grants from Novartis, bioMérieux, Bayer Vital, and Alere and served as consultant or speaker for bioMérieux, Novartis,and Pfizer. JZ received a speaker’s honorarium from Pfizer.Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Background: Many patients admitted to a hospital are already colonized with multi-drug resistant organisms (MDRO) including third-generation cephalosporin-resistant Enterobacteriaceae (3GCREB). The aim of our study was to determine the prevalence of rectal 3GCREB colonization at admission to a large German university hospital and to estimate infection incidences. In addition, risk factors for 3GCREB colonization were identified., Materials/methods: In 2014 and 2015, patients were screened for rectal colonization with 3GCREB and filled out a questionnaire on potential risk factors at admission to a non-intensive care unit (non-ICU). All patients were retrospectively monitored for bacterial infections. Descriptive, univariable and multivariable logistic regression analyses were conducted to identify risk factors for 3GCREB colonization at admission., Results: Of 4,013 patients included, 10.3% (n = 415) were rectally colonized with 3GCREB at admission. Incidence of nosocomial infections was 3.5 (95% CI 2.0-6.1) per 100 patients rectally colonized with 3GCREB compared to 2.3 (95% CI 1.8-3.0, P = 0.213) per 100 3GCREB negative patients. Independent risk factors for 3GCREB colonization were prior colonization / infection with MDRO (OR 2.30, 95% CI 1.59-3.32), prior antimicrobial treatment (OR 1.97, 95% CI 1.59-2.45), male sex (OR 1.38, 95% CI 1.12-1.70), prior travelling outside Europe (OR 2.39, 95% CI 1.77-3.22) and places of residence in the Berlin districts Charlottenburg-Wilmersdorf (OR 1.52, 95% CI 1.06-2.18), Friedrichshain-Kreuzberg (OR 2.32, 95% CI 1.44-3.74) and Mitte (OR 1.73, 95% CI 1.26-2.36)., Conclusions: Admission prevalence of rectal colonization with 3GCREB was high, while infection incidence did not significantly differ between patients rectally colonized or not with 3GCREB at hospital admission. In consequence, hospitals should prioritize improvement of standard precautions including hand hygiene to prevent infections among all patients irrespective of their 3GCREB status at hospital admission., Competing Interests: The authors have declared that no competing interests exist.

Objectives: Fluoroquinolone resistance (FQR) in third-generation cephalosporin-resistant Enterobacteriaceae (3GCRE) presents serious limitations to antibiotic therapy. The aim of this study was to investigate whether the FQR proportion among 3GCRE differs between community-acquired (CA) and hospital-acquired (HA) isolates., Methods: In a prospective observational study covering 2014 and 2015, we monitored the occurrence of 3GCRE in adult hospitalized patients in six German university hospitals. 3GCRE clinical isolates were subdivided into CA and HA. Multivariable analysis identified factors associated with in vitro non-susceptibility to ciprofloxacin., Results: The dataset included 5721 3GCRE isolates of which 52.9% were HA and 52.7% exhibited FQR. Interestingly, the FQR proportion was higher in CA 3GCRE than in HA 3GCRE (overall, 60.1% versus 46.2%, respectively, P < 0.001). Multivariable analysis adjusting for age confirmed community acquisition as a risk factor for FQR [adjusted rate ratio (aRR) 1.33, 95% CI 1.17-1.53]. Escherichia coli and Klebsiella spp. were associated with a much higher FQR proportion than other Enterobacteriaceae species (aRR 8.14, 95% CI 6.86-9.65 and aRR 7.62 with 95% CI 6.74-8.61, respectively)., Conclusions: The high FQR proportion observed among CA 3GCRE, particularly in E. coli and Klebsiella spp., indicates that selection pressure in the outpatient setting needs to be addressed with antibiotic stewardship and other interventions in order to limit further spread of MDR.

A previously healthy 60-year-old patient presented to the emergency department with severe headache, altered personality and fever. He was treated for bacterial meningitis with delirium of unknown cause but presumed to be due to alcohol withdrawal. Despite receiving the antibiotic therapy regimen recommended for bacterial meningitis the patient's condition rapidly deteriorated with profound delirium and tachypnea. The intensivist who was consulted immediately suspected sepsis-associated organ failure and admitted the patient to the intensive care unit (ICU). The blood culture was positive for Listeria. After 10 days the patient could be discharged from the ICU and ultimately recovered completely. In patients presenting with unexplained delirium or altered personality the suspicion of septic encephalopathy should always be considered. They should be admitted to the ICU and sepsis treatment should be initiated without delay.

Background: Bacterial colonisation with Moraxella catarrhalis may partly sustain chronic inflammation in the lower airways of patients with chronic obstructive pulmonary disease (COPD). In addition, this bacterium causes infectious exacerbations of COPD, which often necessitate treatment with antibiotics. Antimicrobial peptides are the body's own antibiotic substances with bactericidal and bacteriostatic, as well as immunomodulatory function. In particular, human beta-defensin 3 (hBD-3) exerts an antimicrobial effect against an extraordinarily broad spectrum of pathogens. We therefore investigated the role of hBD-3 in infections of pulmonary epithelial cells with M. catarrhalis., Methods: The antimicrobial activity of hBD-3 vs. M. catarrhalis was evaluated in an antimicrobial susceptibility assay. We analyzed hBD-3 secretion of M. catarrhalis-infected pulmonary epithelial cells using ELISA. The role of M. catarrhalis-specific virulence factors, toll-like receptors (TLR) 2 and 4, MAPK pathways, and transcription factors AP-1 and NF-κB in the induction and regulation of hBD-3 expression were explored with specific inhibitors, small interference RNA, Western Blot, and chromatin immunoprecipitation (ChIP) assays., Results: HBD-3 exhibited a strong bactericidal effect against M. catarrhalis. M. catarrhalis induced hBD-3 expression in pulmonary epithelial cells, which was dependent on M. catarrhalis membranous lipoolygosaccharide (LOS), while the surface proteins UspA1 and UspA2 were not involved. Gene silencing of TLR2, but not TLR4, led to a reduced hBD-3 secretion after stimulation with M. catarrhalis or M. catarrhalis LOS. Inhibition of MAPKs ERK1/2 and JNK, but not p38, reduced hBD-3 secretion. HBD-3 expression was mediated through the recruitment of AP-1 to the hBD-3 gene promoter and was independent of NF-κB., Conclusion: The immune response of pulmonary epithelial cells towards M. catarrhalis involves secretion of hBD-3, which has a bactericidal effect against this pathogen. Binding of M. catarrhalis virulence factor LOS to TLR2 causes an ERK1/2- and JNK-dependent induction of AP-1-related transcription of the hBD-3 gene, resulting in the production and secretion of hBD-3., (Copyright © 2015. Published by Elsevier Inc.)

Lower respiratory tract bacterial infections are characterized by neutrophilic inflammation in the airways. The carcinoembryonic antigen-related cell adhesion molecule (CEACAM) 8 is expressed in and released by human granulocytes. Our study demonstrates that human granulocytes release CEACAM8 in response to bacterial DNA in a TLR9-dependent manner. Individuals with a high percentage of bronchial lavage fluid (BALF) granulocytes were more likely to have detectable levels of released CEACAM8 in the BALF than those with a normal granulocyte count. Soluble, recombinant CEACAM8-Fc binds to CEACAM1 expressed on human airway epithelium. Application of CEACAM8-Fc to CEACAM1-positive human pulmonary epithelial cells resulted in reduced TLR2-dependent inflammatory responses. These inhibitory effects were accompanied by tyrosine phosphorylation of the immunoreceptor tyrosine-based inhibitory motif (ITIM) of CEACAM1 and by recruitment of the phosphatase SHP-1, which could negatively regulate Toll-like receptor 2-dependent activation of the phosphatidylinositol 3-OH kinase-Akt kinase pathway. Our results suggest a new mechanism by which granulocytes reduce pro-inflammatory immune responses in human airways via secretion of CEACAM8 in neutrophil-driven bacterial infections.

Studying the interaction of dendritic cells (DCs) with bacteria controlled by T-cell-mediated immune responses may reveal novel adjuvants for the induction of cellular immunity. Murine studies and the observation that nocardias infect predominantly immunosuppressed patients have suggested that these bacteria may possess an adjuvant potential. Moreover, adjuvants on the basis of the nocardial cell wall have been applied in clinical studies. Since the handling of adjuvants by DCs may determine the type of immune responses induced by a vaccine, the present study aimed at investigating the interaction of immature human monocyte-derived DCs with live or inactivated Nocardia farcinica in vitro and determining the cellular phenotypic changes as well as alterations in characteristic functions, such as phagocytosis, induction of T-cell proliferation, and cytokine secretion. Human DCs ingested N. farcinica and eradicated the bacterium intracellularly. DCs exposed to inactivated N. farcinica were activated, i.e., they developed a mature phenotype, downregulated their phagocytic capacity, and stimulated allogeneic T cells in mixed leukocyte reactions. Soluble factors were not involved in this process. To elucidate the potential adjuvant effect of N. farcinica on the induction of T-cell-mediated immune responses, we characterized the cytokines produced by nocardia-exposed DCs and detected substantial amounts of tumor necrosis factor alpha (TNF-α) and interleukin-12 p40 (IL-12p40). However, nocardia-treated DCs secreted only small amounts of IL-12p70, which were significantly smaller than the amounts of IL-23. Thus, N. farcinica activates DCs, but adjuvants based on this bacterium may have only a limited capacity to induce Th1 immune responses.

Infections of the central nervous system (CNS) are increasingly reported in patients with malignancies. Heavily immunocompromised patients like those after allogeneic stem cell transplantation (SCT) or previous T cell depleting treatment regimens (e.g. with fludarabine or alemtuzumab) are at highest risk for cerebral infections. The spectrum of causative organisms may vary greatly, depending on the underlying malignancy, its treatment and various other factors. Toxoplasma gondii and fungi are the leading causative organisms in patients after allogeneic SCT, but also viruses such as herpes simplex virus or JC virus may be detected in these patients. Definitive diagnosis of cerebral infection still remains a high challenge, although diagnostics have improved by the wide availability of imaging techniques and polymerase chain reaction in recent years. Novel therapeutic options are arising, particularly for fungal CNS infections. Here, we summarise aspects on epidemiology, clinical symptoms and prognosis of CNS infections in patients with malignancies. Additionally, we give an overview on the diagnostics and management of cerebral infections in these patients including evidence evaluation of efficacy of treatment.

Although Moraxella catarrhalis and Neisseria meningitidis are important human pathogens, they often colonize the human respiratory tract without causing overt clinical symptoms. Both pathogens express structurally unrelated proteins that share the ability to stimulate the adhesion molecule CEACAM1 expressed on human cells. Here we demonstrate that the interaction of CEACAM1 with ubiquitous surface protein A1 expressed on M. catarrhalis or with opacity-associated proteins on N. meningitidis resulted in reduced Toll-like receptor 2-initiated transcription factor NF-kappaB-dependent inflammatory responses of primary pulmonary epithelial cells. These inhibitory effects were mediated by tyrosine phosphorylation of the immunoreceptor tyrosine-based inhibitory motif of CEACAM1 and by recruitment of the phosphatase SHP-1, which negatively regulated Toll-like receptor 2-dependent activation of the phosphatidylinositol 3-OH kinase-Akt kinase pathway. Our results identify a CEACAM1-dependent immune-evasion strategy.

Moraxella catarrhalis is a major cause of exacerbations of chronic obstructive pulmonary disease (COPD) and emphysema. M. catarrhalis-specific UspA1 and the epithelial carcinoembryonic antigen-related cell adhesion molecule (CEACAM1) were required to induce apoptosis. M. catarrhalis-induced apoptosis was significantly enhanced in HeLa cells stably transfected with CEACAM1, compared with HeLa cells not expressing CEACAM1. Infected cells showed increased activity of caspases 3, 6, and 9 but not of caspase 8. Reduced expression of Bcl-2, translocation of Bax into the mitochondria, and cytosolic increase of apoptosis-inducing factor in M. catarrhalis-infected cells implicated the involvement of mitochondrial death pathways. In conclusion, M. catarrhalis induced apoptosis in pulmonary epithelial cells--a process that was triggered by interaction between CEACAM1 and UspA1. Thus, M. catarrhalis-induced apoptosis of pulmonary epithelial cells may contribute to the development of COPD and emphysema.

Reactive oxygen and nitrogen species are released by immune-competent cells and contribute to cellular damage. On the other hand, certain pathogens, including Streptococcus pneumoniae, are known to produce hydrogen peroxide (H2O2), while production of nitrogen radicals by bacteria presumably occurs but has been poorly studied. We determined the relative contributions of bacterial versus host-derived oxygen and nitrogen radicals to cellular damage in pneumococcal infection. A special focus was placed on peroxynitrite as a hypothetical common product formed by the reaction of H2O2 and NO. In microglial cultures, reduction of the formation of 3-nitrotyrosine and cellular damage required H2O2-deficient (DeltaspxB or DeltacarB) pneumococci and inhibition of host NO synthesis with aminoguanidine. In infected C57BL/6 mice, neuronal loss and immunopositivity for nitrotyrosine in the dentate gyrus were markedly reduced with DeltaspxB or DeltacarB bacterial mutants and in inducible nitric oxide synthase knockout mice. We conclude that although host and bacteria both produce oxygen and nitrogen radicals, the interplay of prokaryotic H2O2 and eukaryotic NO is a major contributor to cellular damage in pneumococcal meningitis.

Lipopolysaccharide-binding protein (LBP) has a well-established role in Gram-negative infection. New data suggest a more expanded role for LBP as a general recognition molecule. Several bacterial surface components from Gram-positive pathogens are also recognized by this molecule. LBP may also serve as a clinical marker in severe infections and may carry therapeutic potential.

Streptococcus pneumoniae is the most common cause of bacterial meningitis of high mortality and morbidity. Neurological sequelae include paralysis, mental retardation, and learning disorders. In humans, neurons of the hippocampus undergo apoptosis as a result of meningitis. Phosphatidylcholine (PtdCho) is an essential component of mammalian cell membranes and PtdCho deficiency, either due to chemicals or altered nutrition, leads to apoptosis, especially in hippocampal neurons. We show that apoptosis of a variety of brain cells after pneumococcal infection arises from inhibition of PtdCho biosynthesis, the first such activity described for a bacterium. Apoptosis inhibitors did not prevent the bacterial-dependent inhibition of PtdCho biosynthesis. Supplementation with exogenous lyso-phosphatidylcholine prevents cell death and treatment of mice with cytidine diphosphocholine attenuates hippocampal damage during meningitis, even after the onset of infection. We conclude that bacterial inhibition of PtdCho biosynthesis activates an apoptotic cascade that is a causative event in pathogenesis and amenable to therapeutic intervention.

Doxorubicin induces apoptosis in a variety of cells. We investigated the expression and function of various tumor necrosis factor (TNF)alpha-homologues and their receptors. CEM cells did not differentially express any one of the TNFalpha-homologous receptors investigated nor TNF-related apoptosis-inducing ligand or TNF-related weakly apoptosis-inducing ligand (TWEAK) in the presence of doxorubicin. In addition to CD95 ligand, however, receptor activator of nuclear factor kappaB ligand (RANKL) was strongly up-regulated. Doxorubicin-induced apoptosis was greatly suppressed in the presence of either neutralizing antibody or RANK-Fc fusion protein. Moreover, neutralizing RANKL also prevented cytochrome c release from mitochondria. RANKL alone was unable to induce significant levels of apoptosis in CEM cells. However, doxorubicin-induced apoptosis was increased >2-fold when exogenous RANKL was added. Therefore, RANKL is necessary but not sufficient to account for early doxorubicin-induced apoptosis in CEM cells. This finding suggests improved chemotherapeutic efficiency of the anthracyclin against susceptible malignant cells in the presence with RANKL.

Lipopolysaccharide-binding protein (LBP), an acute-phase protein recognizing lipopolysaccharide (LPS), catalyzes in low concentrations its transfer to the cellular LPS receptor consisting of CD14 and Toll-like receptor-4. It has recently been shown that high concentrations of recombinant LBP can protect mice in a peritonitis model from the lethal effects of LPS. To determine whether in humans the acute-phase rise of LBP concentrations can inhibit LPS binding to monocytes and induction of proinflammatory cytokines, LBP concentrations were analyzed in 63 patients meeting the American College of Chest Physicians/Society of Critical Care Medicine criteria of severe sepsis or septic shock and the ability of these sera to modulate LPS effects in vitro was assessed employing different assays. Transfer of fluorescein isothiocyanate-labeled LPS to human monocytes was assessed by a fluorescence-activated cell sorter-based method, and activation of monocytes was investigated by measuring LPS-induced tumor necrosis factor-alpha secretion in the presence of the sera. Anti-LBP antibodies and recombinant human LBP were instrumental for depletion and reconstitution of acute-phase sera and subsequent assessment of their modulating effects on LPS activity. Sera of patients with severe sepsis/septic shock exhibited a diminished LPS transfer activity and LPS-induced tumor necrosis factor-alpha secretion as compared with sera from healthy controls. LBP depletion of sepsis sera and addition of rhLBP resulting in concentrations found in severe sepsis confirmed that LBP was the major serum component responsible for the observed effects. In summary, the inhibition of LPS effects by high concentrations of LBP in acute-phase serum, as described here, may represent a novel defense mechanism of the host in severe sepsis and during bacterial infections.

Acute phase proteins are extremely helpful markers for indicating a disturbance of the homeostasis within the organism and for monitoring the course of a disease. Despite the availability of several serum acute phase markers, a better and more specific prediction of sepsis and related disorders, such as systemic inflammatory response syndrome (SIRS) is still needed, as these diseases still have a high mortality rate and have to be detected early and with high specificity. Here a novel acute-phase protein is introduced, that has certain biological functions in host defense and that may be a useful addition for the diagnosis and monitoring of sepsis. Lipopolysaccharide (LPS or endotoxin), binding protein (LBP) is a class 1 acute-phase protein with the ability to bind and transfer bacterial LPS. Changes in serum levels of LBP have profound effects on the host's ability to react to endotoxin stimulation and to defend itself against sepsis. Results obtained from in vitro studies and from an animal model are reviewed here and a perspective on ongoing clinical studies is given. There is evidence that LBP, along with other LPS-recognizing molecules, is an important parameter for monitoring the acute phase and the ability of the host to react to LPS-challenge.