OXA-48-like carbapenemases in Proteus mirabilis - novel genetic environments and a challenge for detection.

OXA-48-like enzymes represent the most frequently detected carbapenemases in Enterobacterales in Western Europe, North Africa and the Middle East. In contrast to other species, the presence of OXA-48-like in Proteus mirabilis leads to an unusually susceptible phenotype with low MICs for carbapenems and piperacillin-tazobactam, which is easily missed in the diagnostic laboratory. So far, there is little data available on the genetic environments of the corresponding genes, blaOXA-48-like, in P. mirabilis. In this study susceptibility phenotypes and genomic data of 13 OXA-48-like-producing P. mirabilis were investigated (OXA-48, n=9; OXA-181, n=3; OXA-162, n=1). Ten isolates were susceptible to meropenem and ertapenem and three isolates were susceptible to piperacillin-tazobactam. The gene blaOXA-48 was chromosomally located in 7/9 isolates. Thereof, in three isolates blaOXA-48 was inserted into a P. mirabilis genomic island. Of the three isolates harbouring blaOXA-181 one was located on an IncX3 plasmid and two were located on a novel MOBF plasmid, pOXA-P12, within the new transposon Tn7713. In 5/6 isolates with plasmidic location of blaOXA-48-like, the plasmids could conjugate to E. coli recipients in vitro. Vice versa, blaOXA-48-carrying plasmids could conjugate from other Enterobacterales into a P. mirabilis recipient. These data show a high diversity of blaOXA-48-like genetic environments compared to other Enterobacterales, where genetic environments are quite homogenous. Given the difficult-to-detect phenotype of OXA-48-like-producing P. mirabilis and the location of blaOXA-48-like on mobile genetic elements, it is likely that OXA-48-like-producing P. mirabilis can disseminate, escape most surveillance systems, and contribute to a hidden spread of OXA-48-like.

In Vitro Activity of Two Novel Antimicrobial Compounds on MDR-Resistant Clinical Isolates.

The development of novel antibiotics is mandatory to curb the growing antibiotic resistance problem resulting in difficult-to-treat bacterial infections. Here, we have determined the spectrum of activity of cystobactamids and chelocardins, two novel and promising classes of molecules with different modes of action. A panel of 297 clinically relevant Gram-negative and Gram-positive isolates with different antibiotic susceptibility profiles, going from wild type to multi- or even extremely drug resistant (MDR, XDR) and including carbapenem-resistant isolates, were tested using broth microdilution assays to determine the minimal inhibitory concentrations (MICs), MIC50s and MIC90s of two cystobactamids derivatives (CN-861-2 and CN-DM-861) and two chelocardin derivatives (CHD and CDCHD). Cystobactamids revealed potent activities on the majority of tested Enterobacterales (MIC50s ranging from 0.25 to 4 µg/mL), except for Klebsiella pneumoniae isolates (MIC50s is 128 µg/mL). Pseudomonas aeruginosa and Acinetobacter baumannii showed slightly higher MIC50s (4 µg/mL and 8 µg/mL, respectively) for cystobactamids. Chelocardins inhibited the growth of Enterobacterales and Stenotrophomas maltophilia at low to moderate MICs (0.25-16 µg/mL) and the chemically modified CDCHD was active at lower MICs. A. baumannii and P. aeruginosa were less susceptible to these molecules with MICs ranging from 0.5 to 32 µg/mL. These molecules show also interesting in vitro efficacies on clinically relevant Gram-positive bacteria with MICs of 0.125-8 µg/mL for cystobactamids and 0.5-8 µg/mL for chelocardins. Taken together, the cystobactamid CN-DM-861 and chelocardin CDCHD showed interesting antibiotic activities on MDR or XDR bacteria, without cross-resistance to clinically relevant antibiotics such as carbapenems, fluoroquinolones, and colistin.

Proteus mirabilis - analysis of a concealed source of carbapenemases and development of a diagnostic algorithm for detection.

OBJECTIVES: To analyse carbapenemases in Proteus mirabilis and assess the performance of carbapenemase detection assays. METHODS: Eighty-one clinical P. mirabilis isolates with high-level resistance at least to ampicillin (>32 mg/L) or previous detection of carbapenemases were selected and investigated by three susceptibility testing methods (microdilution, automated susceptibility testing, and disk diffusion), six phenotypic carbapenemase assays (CARBA NP, modified carbapenemase inactivation method [CIM], modified zinc-supplemented CIM, simplified CIM, faropenem, and carbapenem-containing agar), two immunochromatographic assays, and whole-genome sequencing. RESULTS: Carbapenemases were detected in 43 of 81 isolates (OXA-48-like [n = 13]; OXA-23 [n = 12]; OXA-58 [n = 12]; New Delhi metallo-ß-lactamase (NDM) [n = 2]; Verona integron-encoded metallo-ß-lactamase (VIM) [n = 2]; Imipenemase (IMP) [n = 1]; Klebsiella pneumoniae carbapenemase (KPC) [n = 1]). Carbapenemase-producing Proteus were frequently susceptible to ertapenem (26/43; 60%), meropenem (28/43; 65%), ceftazidime (33/43; 77%), and some even to piperacillin-tazobactam (9/43; 21%). Sensitivity/specificity of phenotypic tests were 30% (CI: 17-46%)/89% (CI: 75-97%) for CARBA NP, 74% (CI: 60-85%)/82% (CI: 67-91%) for faropenem, 91% (CI: 78-97%)/82% (CI: 66-92%) for simplified CIM, and 93% (CI: 81-99%)/100% (CI: 91-100%) for modified zinc-supplemented CIM. An algorithm for improved detection was developed, which demonstrated sensitivity/specificity of 100% (CI: 92-100%)/100% (CI: 91-100%) on the 81 isolates, and 100% (CI: 29-100%)/100% (CI: 96-100%) in a prospective analysis of additional 91 isolates. Interestingly, several OXA-23-producing isolates belonged to the same clonal lineage reported previously from France. DISCUSSION: Current susceptibility testing methods and phenotypic tests frequently fail to detect carbapenemases in P. mirabilis, which could result in inadequate antibiotic treatment. In addition, the non-inclusion of blaOXA-23/OXA-58 in many molecular carbapenemase assays further impedes their detection. Therefore, the prevalence of carbapenemases in P. mirabilis is likely underestimated. With the herein proposed algorithm, carbapenemase-producing Proteus can be easily identified.

An appeal for strengthening genomic pathogen surveillance to improve pandemic preparedness and infection prevention: the German perspective.

The SARS-CoV-2 pandemic has highlighted the importance of viable infection surveillance and the relevant infrastructure. From a German perspective, an integral part of this infrastructure, genomic pathogen sequencing, was at best fragmentary and stretched to its limits due to the lack or inefficient use of equipment, human resources, data management and coordination. The experience in other countries has shown that the rate of sequenced positive samples and linkage of genomic and epidemiological data (person, place, time) represent important factors for a successful application of genomic pathogen surveillance. Planning, establishing and consistently supporting adequate structures for genomic pathogen surveillance will be crucial to identify and combat future pandemics as well as other challenges in infectious diseases such as multi-drug resistant bacteria and healthcare-associated infections. Therefore, the authors propose a multifaceted and coordinated process for the definition of procedural, legal and technical standards for comprehensive genomic pathogen surveillance in Germany, covering the areas of genomic sequencing, data collection and data linkage, as well as target pathogens. A comparative analysis of the structures established in Germany and in other countries is applied. This proposal aims to better tackle epi- and pandemics to come and take action from the "lessons learned" from the SARS-CoV-2 pandemic.

In Vitro Activity of Cefiderocol against Clinical Gram-Negative Isolates Originating from Germany in 2016/17.

Antimicrobial resistance poses a global threat to public health. Of great concern are Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacterales with resistance to carbapenems or third-generation cephalosporins. The aim of the present study was to investigate the in vitro activity of the novel siderophore cephaloporin cefiderocol (CID) and four comparator ß-lactam-ß-lactamase-inhibitor combinations and to give insights into the genetic background of CID-resistant isolates. In total, 301 clinical Enterobacterales and non-fermenting bacterial isolates were selected for this study, including randomly chosen isolates (set I, n = 195) and challenge isolates (set II, n = 106; enriched with ESBL and carbapenemase producers, as well as colistin-resistant isolates). Isolates displayed CID MIC50/90 values of 0.12/0.5 mg/L (set I) and 0.5/1 mg/L (set II). Overall, the CID activity was superior to the comparators against A. baumannii, Stenotrophomonas maltophilia and set II isolates of P. aeruginosa. There were eight CID-resistant isolates detected (MIC > 2 mg/L): A. baumannii (n = 1), E. cloacae complex (n = 5) and P. aeruginosa (n = 2). Sequencing analyses of these isolates detected the acquired ß-lactamase (bla) genes blaNDM-1,blaSHV-12 and naturally occurring blaOXA-396, blaACT-type and blaCMH-3. In conclusion, CID revealed potent activity against clinically relevant organisms of multidrug-resistant Enterobacterales and non-fermenters.

Susceptibility of Multidrug-Resistant Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa from Germany to Ceftolozane-Tazobactam, Ceftazidime-Avibactam, and Imipenem-Relebactam.

This cross-sectional in-vitro resistance surveillance study involving 10 medical laboratories was conducted in 2018. Each study site was asked to collect 30 consecutive nonduplicate isolates per species from hospitalized patients with documented infections. Minimum inhibitory concentrations were determined at a central laboratory. European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints were used for interpretation. A total of 860 isolates were collected, including 298 Escherichia coli, 268 Klebsiella pneumoniae, and 294 Pseudomonas aeruginosa. Fifty (16.8%) E. coli and 63 (23.5%) K. pneumoniae isolates were found to be resistant to third-generation cephalosporins. Resistance to carbapenems (imipenem and/or meropenem) was identified in 5 (1.9%) K. pneumoniae and 64 (21.8%) P. aeruginosa, but not in E. coli. Thirty-three (11.2%) P. aeruginosa isolates were resistant to both carbapenems and 30 (10.2%) P. aeruginosa showed resistance to ≥3 antimicrobials/antimicrobial groups (among piperacillin-tazobactam, ceftazidime, tobramycin, carbapenems, and fluoroquinolones). The susceptibility rates of these multidrug-resistant (MDR) phenotypes to ceftolozane-tazobactam, ceftazidime-avibactam, and imipenem-relebactam were 70-100%, with the exception of carbapenem-resistant K. pneumoniae. Only two K. pneumoniae and four P. aeruginosa isolates were resistant to all three beta-lactam/beta-lactamase-inhibitor combinations. However, this favorable result should be viewed in light of the relatively low prevalence of MDR organisms that require these agents in Germany.

Increased zinc levels facilitate phenotypic detection of ceftazidime-avibactam resistance in metallo-ß-lactamase-producing Gram-negative bacteria.

Ceftazidime-avibactam is one of the last resort antimicrobial agents for the treatment of carbapenem-resistant, Gram-negative bacteria. Metallo-ß-lactamase-producing bacteria are considered to be ceftazidime-avibactam resistant. Here, we evaluated a semi-automated antimicrobial susceptibility testing system regarding its capability to detect phenotypic ceftazidime-avibactam resistance in 176 carbapenem-resistant, metallo-ß-lactamase-producing Enterobacterales and Pseudomonas aeruginosa isolates. Nine clinical isolates displayed ceftazidime-avibactam susceptibility in the semi-automated system and six of these isolates were susceptible by broth microdilution, too. In all nine isolates, metallo-ß-lactamase-mediated hydrolytic activity was demonstrated with the EDTA-modified carbapenemase inactivation method. As zinc is known to be an important co-factor for metallo-ß-lactamase activity, test media of the semi-automated antimicrobial susceptibility testing system and broth microdilution were supplemented with zinc. Thereby, the detection of phenotypic resistance was improved in the semi-automated system and in broth microdilution. Currently, ceftazidime-avibactam is not approved as treatment option for infections by metallo-ß-lactamase-producing, Gram-negative bacteria. In infections caused by carbapenem-resistant Gram-negatives, we therefore recommend to rule out the presence of metallo-ß-lactamases with additional methods before initiating ceftazidime-avibactam treatment.

Detection of OXA-181-producing Pseudomonas aeruginosa in Germany.

OBJECTIVES: To report the detection of the class D carbapenemase OXA-181 in an MDR clinical Pseudomonas aeruginosa isolate in Germany. METHODS: Carbapenemase detection was performed by using several phenotypic tests such as the modified Hodge test, a combined disc test with boronic acid, EDTA or cloxacillin, a lysate-based inhibition assays and by PCR for common and rare carbapenemase genes. Antibiotic susceptibilities were determined by broth microdilution. The genetic environment of blaOXA-181 in the clinical P. aeruginosa isolate was characterised by Illumina and MinION sequencing. RESULTS: An multidrug-resistant P. aeruginosa was isolated from a tracheal swab in 2019 and was sent to the German National Reference Centre for multidrug-resistant Gram-negative bacteria for carbapenemase detection. Several phenotypic tests indicated the presence of a carbapenemase which was not inhibited by EDTA nor by boronic acid. PCRs for common and rare carbapenemase genes revealed the presence of a blaOXA-181 gene. WGS data confirmed that the gene was located on the chromosome as part of a Tn2013 transposon. The genetic organisation of blaOXA-181 has already been described in a P. aeruginosa isolate from England, but both isolates differed significantly in their sequence types (ST111/ST235). Analysis of the genetic environment of the blaOXA-181 gene also revealed high homology to a plasmid from a Klebsiella pneumoniae isolate. CONCLUSIONS: To our knowledge, this is the first report of blaOXA-181 in a clinical P. aeruginosa isolate in Germany which emphasises the ongoing spread of yet unusual carbapenemases among different Gram-negative species and therefore complicating their detection in routine laboratories.

Effect of sigma E on carbapenem resistance in OXA-48-producing Klebsiella pneumoniae.

OBJECTIVES: Resistance levels of Gram-negative bacteria producing OXA-48 carbapenemase can vary greatly and some of them can even be categorized as susceptible to imipenem and meropenem according to EUCAST breakpoints. This study aimed to reveal resistance mechanisms leading to varying levels of resistance to carbapenems in Klebsiella pneumoniae with blaOXA-48 submitted to the German National Reference Centre for MDR Gram-negative bacteria. METHODS: Meropenem-susceptible clinical blaOXA-48-bearing K. pneumoniae isolates were put under gradually increasing selective pressure of meropenem. Clinical isolates and spontaneous meropenem-resistant mutants were whole-genome sequenced with Illumina and Oxford Nanopore Technology. Identified mutations apart from porin mutations were genetically constructed in the original clinical isolates using CRISPR/Cas. Clinical isolates and mutants were analysed for MICs, growth rates and expression of porins on mRNA and protein levels. RESULTS: Mutations associated with meropenem resistance were predominantly found in ompK36, but in some cases ompK36 was unaffected. In two mutants, ISs within the rpoE (sigma factor E; σE) operon were detected, directly in or upstream of rseA. These IS1R elements were then inserted into the same position of the susceptible clinical isolates using CRISPR/Cas. CRISPR-rseA-rseB-rseC mutants showed higher resistance levels to carbapenems and cephalosporins, reduced growth rates and reduced expression of major porins OmpK36 and OmpK35 in quantitative RT-PCR and SDS-PAGE. CONCLUSIONS: Enhanced synthesis of σE leads to increased resistance to cephalosporins and carbapenems in clinical K. pneumoniae isolates. This effect could be based upon remodelling of expression patterns of outer membrane proteins. The up-regulated σE stress response also leads to a significant reduction in growth rates.

Characterization of GMB-1, a novel metallo-ß-lactamase (MBL) found in three different Enterobacterales species.

OBJECTIVES: To identify novel carbapenem resistance mechanisms and their potential to spread among clinical isolates. METHODS: Four clinical isolates of Citrobacter freundii, Serratia marcescens and Raoultella planticola (n = 2) from one hospital in Central Germany were sent to the German National Reference Centre for Multidrug-resistant Gram-negative Bacteria for carbapenemase detection. Phenotypic tests indicated the presence of a metallo-ß-lactamase (MBL), but PCR for various MBL genes could not identify any. Using WGS data, a putative bla gene was identified. Its carbapenemase activity was verified by heterologous expression in an Escherichia coli cloning strain, with subsequent MIC determination by broth microdilution, as well as by in vitro hydrolysis assays using purified enzyme. RESULTS: WGS indicated the presence of a putative ß-lactamase with 48% amino acid identity to the subclass B1 MBL SPM-1. MIC studies confirmed that the novel enzyme formed a functional MBL, which was therefore designated as GMB-1 (German MBL). In vitro hydrolysis assays showed a lack of activity not only against aztreonam but also against ertapenem. WGS revealed that in all three species the blaGMB-1 gene was located on the chromosome as part of a genetic island with multiple ISs. CONCLUSIONS: The finding of GMB-1 once again shows that novel carbapenemases continue to emerge and make their way into clinically relevant species. The occurrence of GMB-1 in three different species demonstrates the extraordinary mobility of such genetic islands and their potential to spread carbapenemase genes into diverse genetic environments.

Seroprevalence of SARS-CoV-2 Antibodies in Employees of Three Hospitals of a Secondary Care Hospital Network in Germany and an Associated Fire Brigade: Results of a Repeated Cross-Sectional Surveillance Study Over 1 Year.

Healthcare workers (HCWs) are playing a vital role in the current SARS-CoV-2 pandemic. This study investigated how infection spreads within three local hospitals and an associated fire brigade in Germany by testing employees for the presence of SARS-CoV-2 IgG antibodies over one year. The three observational periods corresponded to the initial three pandemic waves: first wave: June-September 2020, second wave: October 2020-January 2021, and third wave: February-June 2021. We analysed 3285 serum samples of 1842 employees, which represents 65.7% of all employees. Altogether, 13.2% employees were seropositive: 194/1411 HCWs (13.7%) and 49/431 non-HCWs (11.4%) with a clear increase of seroprevalence from the first (1.1%) to the second (13.2%) and third (29.3%) pandemic wave. HCWs presumably had an additional occupational risk for infection in the second and third wave due to an increase of infection pressure with more COVID-19 patients treated, showing possible weak points in the recommended infection prevention strategy.

Geographical differences of carbapenem non-susceptible Enterobacterales and Acinetobacter spp. in Germany from 2017 to 2019.

BACKGROUND: Since May 2016, infection and colonisation with carbapenem non-susceptible Acinetobacter spp. (CRA) and Enterobacterales (CRE) have to be notified to health authorities in Germany. The aim of our study was to assess the epidemiology of CRA and CRE from 2017 to 2019 in Germany, to identify risk groups and to determine geographical differences of CRA and CRE notifications. METHODS: Cases were notified from laboratories to local public health authorities and forwarded to state and national level. Non-susceptibility was defined as intermediate or resistant to ertapenem, imipenem, or meropenem excluding intrinsic bacterial resistance or the detection of a carbapenemase gene. We analysed CRA and CRE notifications from 2017, 2018 and 2019 per 100,000 inhabitants (notification incidence), regarding their demographic, clinical and laboratory information. The effect of regional hospital-density on CRA and CRE notification incidence was estimated using negative binomial regression. RESULTS: From 2017 to 2019, 2278 CRA and 12,282 CRE cases were notified in Germany. CRA and CRE cases did not differ regarding demographic and clinical information, e.g. proportion infected. The notification incidence of CRA declined slightly from 0.95 in 2017 to 0.86 in 2019, whereas CRE increased from 4.23 in 2017 to 5.72 in 2019. The highest CRA and CRE notification incidences were found in the age groups above 70 years. Infants below 1 year showed a high CRE notification incidence, too. Notification incidences varied between 0.10 and 2.86 for CRA and between 1.49 and 9.99 for CRE by federal state. The notification incidence of CRA and CRE cases increased with each additional hospital per district. CONCLUSION: The notification incidence of CRA and CRE varied geographically and was correlated with the number of hospitals.The results support the assumption that hospitals are the main driver for higher CRE and CRA incidence. Preventive strategies and early control measures should target older age groups and newborns and areas with a high incidence.

Evaluation of the rCIM for carbapenemase detection in Enterobacterales and Pseudomonas aeruginosa and description of the TSBrCIM, an optimized variant.

OBJECTIVES: The rapid Carbapenem Inactivation Method (rCIM) was evaluated with a strain collection of 164 and 69 carbapenem-resistant Enterobacterales and Pseudomonas aeruginosa, respectively, that produced various carbapenemases. For an improved carbapenemase detection in Enterobacterales, an optimized variant of the rCIM named TSBrCIM was developed. METHODS: Bacterial isolates were incubated with two meropenem disks in distilled water (rCIM) or tryptic soy broth (TSBrCIM). After centrifugation, the supernatant was incubated with a susceptible E. coli indicator strain in tryptic soy broth. Growth of the indicator strain implied carbapenemase activity in the test strain. RESULTS: The rCIM detected 100/113 carbapenemase-producing Enterobacterales, resulting in a sensitivity of 88.5% and a specificity of 94.1%. For P. aeruginosa, sensitivity and specificity were 96.0% and 100%, respectively. The TSBrCIM was able to detect 105/113 carbapenemase-producing Enterobacterales, resulting in a sensitivity of 92.9% and a specificity of 96.1%. CONCLUSION: This study shows that the TSBrCIM can be valuable tool for detection of carbapenemases in Enterobacterales in the clinical laboratory, while the rCIM showed the best results for carbapenemase detection in P. aeruginosa.

COVID-19: Hotspot hospital?- seroprevalence of SARS-CoV-2 antibodies in hospital employees in a secondary care hospital network in Germany: Intermediate results of a prospective surveillance study.

PURPOSE: The objective of the ongoing study was to investigate how SARS-CoV-2 infection spread within two hospitals in North Rhine-Westphalia, Germany by testing the employees working in high-risk, intermediate-risk and low-risk-areas for the presence of SARS-CoV-2 IgG antibodies. Presented intermediate results evaluate the first infection period until the end of September 2020. METHODS: The study "COVID-19: Hotspot hospital?- Seroprevalence of SARS-CoV-2 antibodies in hospital employees in a secondary care hospital network in Germany " is a prospective, single centre observational cohort study conducted at the St. Vincenz Hospital Datteln with 316 beds. The presented data include one other hospital: St. Laurentius Stift Waltrop, Germany with 172 beds. RESULTS: Between June 2020 and September 2020 we analyzed serum samples of 907 employees which represents 62.1% of all employees. Thirteen employees (1.4%), respectively 13/696 healthcare workers (HCWs) (1.9%) had detectable SARS-CoV-2 IgG antibodies. Among them, 4 (30.8%) were aware of COVID-19 exposure, and 5 (38.5%) reported clinical symptoms. HCWs working in high-risk areas had a seroprevalence rate of 1.6% (1/64), HCWs working in intermediate-risk area 1.7% (11/632) and 0.5% employees (1/211) in low-risk areas with no contact to patients were seropositive. CONCLUSION: Even if we treated COVID-19 positive patients, we found no clear evidence that infection was transmitted to HCWs in contact to these patients. As knowledge about SARS-CoV-2 transmission evolves, the concept of infection prevention must be continuously reviewed and adapted as needed to keep hospitals a safe place.

Phenotypic Detection and Differentiation of Carbapenemase Classes Including OXA-48-Like Enzymes in Enterobacterales and Pseudomonas aeruginosa by a Highly Specialized Micronaut-S Microdilution Assay.

The objective of this study was to evaluate the Micronaut-S carbapenemase detection microtiter plate assay for the detection of carbapenemases and Ambler class determination. The Micronaut-S carbapenemase detection microtiter plate was tested using a challenging collection of 154 carbapenemase-producing and 150 carbapenemase-negative clinical strains of Enterobacterales and Pseudomonas aeruginosa The Micronaut-S carbapenemase detection assay was able to detect 148/154 carbapenemase producers correctly, whereas 5/150 non-carbapenemase-producing isolates tested as false positive. This resulted in an overall sensitivity of 96% and a specificity of 97%. Regarding the detection of the carbapenemase class, the sensitivities and specificities were 93%/100%, 96%/100%, and 97%/99% for class A (n = 27), class B (n = 54), and class D (n = 73) carbapenemases, respectively. The Micronaut-S carbapenemase detection microtiter plate represents an easy-to-use and valuable tool for accurate and reliable detection of carbapenemases. In addition, it provides identification of the class of carbapenemase in most cases which can provide significant therapy guidance.

In vitro activity of cefiderocol against aerobic Gram-negative bacterial pathogens from Germany.

OBJECTIVES: Cefiderocol (CID), also known as S-649266, a novel siderophore cephalosporin, possesses potent activity against multidrug-resistant aerobic Gram-negative bacteria (GNB). This study aimed to determine the in vitro activity of CID against two different sets of GNB: i) a random sample of 213 clinical isolates, including 17 extended-spectrum beta-lactamase (ESBL) producers, obtained from intensive care unit patients with nosocomial infections collected during a multicentre surveillance study (set I); and ii) a group of 59 challenge GNB producing various types of carbapenemases (CP; set II). METHODS: Minimum inhibitory concentrations (MICs) were determined using the microdilution method according to the standard ISO 20776-1. Iron-depleted medium was used for testing CID. RESULTS: CID inhibited 97.2% of set I isolates at the EUCAST susceptibility breakpoint of ≤ 2 mg/L. The concentrations of CID inhibiting 50% and 90% (MIC50/90) of the Enterobacterales isolates (n = 146) were 0.12/1.0 mg/L, with ESBL-positive isolates tending to exhibit higher MICs than ESBL-negative isolates to CID. MIC50/90 values of CID for isolates of the Acinetobacter baumannii group (n = 13) and Pseudomonas aeruginosa (n = 54) were 0.06/0.12 mg/L and 0.12/0.5 mg/L, respectively. Further, CID inhibited 88.1% of set II CP-producing isolates at ≤ 2 mg/L. All seven class D CP-producing Acinetobacter baumannii were inhibited at ≤ 0.25 mg/L. MIC50/90 values for CP-producing Enterobacterales (n = 30) and Pseudomonas aeruginosa (n = 22) were 1/4 mg/L and 0.5/2 mg/L, respectively. CONCLUSION: CID showed potent activity against Acinetobacter baumannii, Enterobacterales and Pseudomonas aeruginosa, including CP-producing isolates. Overall, CID inhibited 259 of 272 (95.2%) GNB at ≤ 2 mg/L.

New EUCAST definitions of S, I and R from 2019 - German physicians are largely not aware of the changes.

PURPOSE: On January 1st 2019, the new EUCAST definitions of susceptibility testing categories S, I and R took effect. The changes in the I category have considerable clinical impact because they lead to major changes in the antibiogram, and misinterpretation may result in inappropriate selection and dosing of antibiotics hampering effective treatment of infectious diseases. We assessed if German physicians are aware of the new definitions and their consequences. METHODS: We conducted a nationwide web-based survey to assess the knowledge on the new definitions of S, I and R. The survey was addressed to clinicians across all medical specialties working in Germany and was open from May 9th to July 30th 2019. RESULTS: The answers of 902 participants were included in the analysis. Most participants were employed at hospitals (79.3%) and had already completed specialist training (86.1%). The predominant specialty was internal medicine (50.6%). Of all participants, 45.7% did not know that there was a change in the definitions of S, I and R, and 65.4% did not feel well-informed about the changes. When the participants had to identify true and false statements regarding the new I, substantial knowledge gaps were apparent. Worst results were achieved by those physicians who are not employed in a hospital but work in their own practice. CONCLUSION: Our survey shows that German physicians are insufficiently informed about the new definitions of S, I and R. Further education is strongly needed to ensure optimal treatment of infectious diseases.

Dissemination of carbapenem-resistant Pseudomonas aeruginosa isolates and their susceptibilities to ceftolozane-tazobactam in Germany.

Pseudomonas aeruginosa (PA) is a major cause of healthcare-associated infections. Antipseudomonal carbapenems are among the antimicrobial agents used to treat PA infections, but several mechanisms of resistance, including the production of a carbapenemase (CP), may compromise their clinical efficacy. The objectives of this study were to determine: (i) the dissemination of carbapenem-resistant CP-negative and CP-positive PA isolates; and (ii) the in-vitro activity of ceftolozane-tazobactam (CTT) against carbapenem-susceptible and carbapenem-resistant isolates. Isolates were collected prospectively from January 2016 to April 2017 at 20 German medical laboratories. Each centre was asked to provide 50 consecutive isolates from hospitalized patients. Overall, 985 isolates were collected, of which 34% were obtained from intensive care patients. Seven hundred and thirty-eight (74.9%) isolates were susceptible to both imipenem and meropenem (Subgroup I), and 247 (25.1%) isolates were resistant to carbapenems (Subgroup II): 125 (12.7%) were imipenem-resistant but meropenem-susceptible, 12 (1.2%) were meropenem-resistant but imipenem-susceptible, and 110 (11.2%) were resistant to both carbapenems (Subgroup III). A CP was detected in 28 (2.8%) isolates (predominantly VIM-2). Nine hundred and fifty (96.4%) isolates were CTT-susceptible. Susceptibility to CTT was seen in 99.6% of Subgroup I isolates, 87% of Subgroup II isolates and 74.5% of Subgroup III isolates. Overall, 2.8% of PA produced a CP, while 22.2% were carbapenem-resistant, CP-non-producing isolates. Based on these findings, CTT may be considered for treatment of PA infections, particularly those caused by multi-drug-resistant CP-non-producing isolates.

Propionic Acid Shapes the Multiple Sclerosis Disease Course by an Immunomodulatory Mechanism.

Short-chain fatty acids are processed from indigestible dietary fibers by gut bacteria and have immunomodulatory properties. Here, we investigate propionic acid (PA) in multiple sclerosis (MS), an autoimmune and neurodegenerative disease. Serum and feces of subjects with MS exhibited significantly reduced PA amounts compared with controls, particularly after the first relapse. In a proof-of-concept study, we supplemented PA to therapy-naive MS patients and as an add-on to MS immunotherapy. After 2 weeks of PA intake, we observed a significant and sustained increase of functionally competent regulatory T (Treg) cells, whereas Th1 and Th17 cells decreased significantly. Post-hoc analyses revealed a reduced annual relapse rate, disability stabilization, and reduced brain atrophy after 3 years of PA intake. Functional microbiome analysis revealed increased expression of Treg-cell-inducing genes in the intestine after PA intake. Furthermore, PA normalized Treg cell mitochondrial function and morphology in MS. Our findings suggest that PA can serve as a potent immunomodulatory supplement to MS drugs.