7th joint conference "Joint Microbiology & Infection Conference 2024" of the DGHM and VAAM

Professor

Chair

Antimicrobial resistance surveillance of invasive Bacteroides fragilis isolated from blood, Europe, 2022

Diagnostic and Clinical Microbiology

QuestionBacteroides fragilis is the most frequent cause of anaerobic bacteraemia. Although recent data suggest a rise in resistance to antibiotics commonly recommended for therapy of this and other anaerobic bacteria, surveillance on antimicrobial resistance (AMR) remains limited due to a lack of both data availability and comparability. However, a newly introduced standardised method for antimicrobial susceptibility testing (AST) of anaerobic bacteria has made larger scale surveillance possible for the first time. Thus, a European multicentre study was initiated to investigate phenotypic AMR of Bacteroides fragilis isolates from bacteraemia across Europe in 2022.MethodsClinical microbiology laboratories in Europe were invited to contribute results of antimicrobial susceptibility testing (AST) for blood culture isolates identified as Bacteroides fragilis (per unique patient per year of observation (2022)). Antimicrobial susceptibility testing was performed locally by all participating laboratories in a prospective or retrospective way, using the EUCAST disc diffusion method on Fastidious Anaerobe Agar with 5% mechanically defibrinated horse blood (FAA-HB).ResultsA total of 16 European countries (14 EU plus Switzerland and Israel) reported 449 unique isolates of Bacteroides fragilis from blood for which AST had been performed (out of a total 602 unique isolates). Clindamycin showed the highest resistance rates (20.9%, range 0 - 63.6%), followed by piperacillin-tazobactam (11.1%, 0 - 54.5%), meropenem (13.4%, 0 - 45.5%), and metronidazole (3.1%, 0 - 40.0%), all with wide variation between countries.ConclusionsConsidering the regional differences (unexpectedly high resistance rates in some countries particularly in Southern Europe) and the mean resistance rates across Europe (above 10% for three of the four anti-anaerobic antibiotics included), both local AST of clinically relevant isolates of Bacteroides fragilis and continued surveillance on an international level is warranted.

Oral Presentation

Prof. Dr.

Assessing mecillinam resistance in E. coli strains causing UTI: A comparative evaluation of phenotypic testing methods

Question Mecillinam is recommended by German guidelines as a first-line treatment for uncomplicated urinary tract infections (uUTI). However, comprehensive data on antibiotic susceptibility are lacking due to the rare demand for microbiological diagnostics for uUTIs as well as the challenge of performing susceptibility testing with agar dilution as the reference method. We aim to evaluate various phenotypic methods for reliably predicting mecillinam resistance in E. coli, the most common UTI pathogen.Methods 40 E. coli strains from routine urine cultures were selected, including 20 randomly chosen and 20 with elevated or resistant MIC to mecillinam based on Vitek testing with Ast-N371 (BioMérieux). 5 methods were compared in triplicates: agar dilution, Vitek Ast-N371, agar gradient diffusion (AGD), disk diffusion, and broth microdilution conducted in two different media—Müller-Hinton Broth (MHB) and artificial urine (AU). Categorial agreement was assessed based on the median, using EUCAST susceptibility breakpoints (MIC ≤8 mg/L, zone diameter ≥15 mm). Major errors (ME) were defined as false resistance and very major errors (VME) as false susceptibility compared to the reference method.Results Agar dilution categorized 28 strains as susceptible and 12 as resistant, with 6 near the breakpoint (8 mg/L or 16 mg/L). Categorial agreement to the reference method was 90% for disk diffusion, 83% for Vitek, 78% for AGD, 63% for MHB and 80% for AU. AGD had the most VME (n = 10), generally underestimating MIC values, while disk diffusion was the most reliable method with no ME and 4 VME. Notably, 3 of the 4 VME resulted in strains with an MIC of 16 mg/L. Microdilution methods, including Vitek, generally demonstrated a low reliability, with decreased reproducibility and more ME/ VME compared to disk diffusion. Only 2/12 resistant strains and 13/28 susceptible strains showed categorial agreement in all employed methods.Conclusion Mecillinam resistance testing remains challenging, with routine methods yielding unreliable results. The next phase of the study aims to elucidate the underlying genes associated with mecillinam resistance.

Epidemiology of carbapenemases in Morganella spp., Serratia spp. and Providencia spp.

IntroductionMorganella spp., Serratia spp. and Providencia spp. (MSP) are Enterobacterales that cause infections of the urinary tract, wounds and bloodstream among others. Currently, the epidemiology of carbapenemases in MSP is largely unknown.GoalsTo determine the epidemiology of carbapenemases in MSP and performance of diagnostic assays.Materials &amp; MethodsIn total, 131 clinical carbapenemase-producing MSP patient isolates were included (Serratia marcescens, n=107; Morganella morganii, n=13; Providencia spp., n=11). MSP were isolated from clinical samples at the University Hospitals Cologne, Frankfurt and the German National Reference Centre for multidrug-resistant Gram-negative bacteria. Susceptibility was determined by broth microdilution (BMD). Carbapenemase production was assessed by modified zinc-supplemented carbapenem inactivation method (mzCIM). Growth on carbapenem agar (mSuperCarba) was additionally determined. Resistance genes were analysed by whole genome sequencing.ResultsOXA-48-like were the most frequent carbapenemases (n=112; 85.5%), followed by KPC (n=10; 7.6%), IMP (n=6; 4.5%) and GES (n=3; 2.3%). KPC and GES were only detected in S. marcescens. Interestingly, only 29 isolates (22.1%) co-produced an ESBL, with CTX-M-15 being the most frequent (n=21). In contrast, genes coding for AmpC beta-lactamases were detected in 92 isolates (70.2%). Despite carbapenemase production, susceptibility (S/I) was observed for piperacillin-tazobactam in one isolate, ceftazidime in 61, ertapenem in 10 and meropenem in 42 isolates. All isolates were correctly identified as carbapenemase-producers by mzCIM, but only 120 grew on the selective carbapenem agar.SummaryCarbapenemase production in MSP was mostly observed in S. marcescens and OXA-48-like were by far the most common carbapenemases. Further carbapenemases and ESBL production were less frequent compared to other Enterobacterales species. Despite carbapenemase production, MSP isolates frequently tested susceptible for ceftazidime and meropenem, which could result in insufficient detection in the routine microbiology laboratory.

Dr. med.

Discrimination of Acinetobacter baumannii from Acinetobacter nosocomialis using lipid extractions and subsequent rapid MALDI-TOF MS analysis in negative ion mode

Introduction: Precise species identification is crucial, especially for infection control. Due to its accuracy, MALDI-TOF MS has become a standard identification method in many microbiology laboratories. Nevertheless, for species with very similar proteomic spectra differentiation capability is limited, e.g. between Acinetobacter baumannii and A. nosocomialis. Using MALDI in negative ion mode with lipids as target might overcome this limitation.Goals: Here, we investigated lipid extracts for the discrimination of A. baumannii from A. nosocomialis using a MALDI-TOF MS-based rapid workflow in negative ion mode.Materials &amp; Methods: Heat-inactivated isolates of genetically characterized A. baumannii (n=30) and A. nosocomialis (n=25) including each type strain, were processed with the MBT Lipid Xtract Kit (Bruker Daltonics, Bremen, Germany) MALDI spectra of the obtained lipid extracts were acquired with an MBT Sirius mass spectrometer (Bruker Daltonics) in negative ion mode and analysed using FlexAnalysis software.Results: The lipid spectra of A. baumannii and A. nosocomialis reproducibly showed each specific peaks at m/z 1728, 1881 and 1910 with different intensities, corresponding to lipid A molecules of the outer membrane. A. baumannii showed high intensities for the two peaks at m/z 1728 and 1910 while A. nosocomialis showed high intensities at m/z 1881 instead. Three specific peaks at m/z 2004, 2018 and 2032 were only present in A. nosocomialis spectra, which could be used for species discrimination (Fig. 1).In total, 55 spectra were analyzed. The discriminating peaks showed a consistent appearance amongst the different strains of both species. The presence of the peaks at m/z 2004, 2018 and 2032 and simultaneously high intensities of the peak at m/z 1881 discriminated A. nosocomialis from A. baumannii. In total 54/55 (98%) strains were correctly identified using these specific lipid markers.Summary: Differentiating lipid markers could be used in a decision tree analysis to discriminate the two species A. baumannii and A. nosocomialis reliably and quickly. Further studies will have to be carried out to establish an algorithm for clinical diagnostics.

Abstract: Wastewater-based epidemiology (WBE) can serve and support the public health sector by providing additional valuable indicators of infection incidence in communities. Though it was first used over 80 years ago to monitor Polio outbreaks, WBE experienced a resurgence after the onset of the SARS-CoV 2 pandemic, with global research efforts and collaborations improving sampling infrastructure and developing new analysis methods. Wastewater monitoring of SARS-CoV-2 has been shown to provide early warnings of increased incidence in communities and uncover variant occurrence profiles across time and geographic space. We have been able to expand the horizon of wastewater surveillance by leveraging the now well-established WBE infrastructures to monitor trends in the spread of other pathogens, including monkeypox and influenza. Due to the modular nature of wastewater infrastructure, surveying the emergence of many diseases and mutations is possible at multiple community levels (buildings, neighbourhoods, cities, airports, and individual flights), allowing for tailored public health responses. Next-generation sequencing of wastewater allows for whole or near whole genome resolution of the genetic assemblies of circulating bacterial and viral pathogens and antimicrobial resistance genes, providing a holistic understanding of pathogen dynamics within a community. Since the beginning of the COVID-19 pandemic, working with health authorities, we have been monitoring SARS-CoV-2 in wastewater at the National and European level in our lab, which has allowed us to observe the emergence of SARS-CoV-2 variants in wastewater shed region before reported in clinical data. We have also been successfully monitoring SARS-CoV-2 variants on incoming flights to the Frankfurt International Airport, as well as profiling variants in the surrounding urban communities. We are also looking at the impact of COVID-19 on circulating antimicrobial resistance, as it will provide information about whether the change in people's behaviour toward disinfectants and hygiene practices impacted antimicrobial resistance. We are expanding our methods to profile other diseases in wastewater to further utilize the potential of WBE as an early monitoring system as a part of the One-Health concept.

Wastewater Based Epidemiology – Potentials for pandemic management

Invited Speaker

Invited Talk

Shine like a diamond: Rapid ATP measurements to evaluate the efficacy of different cleaning methods for worn spectacles.

Healthcare-associated infections and pathogens: Prevention, surveillance, outbreaks und antibiotic stewardship

Spectacles can harbor various types of microorganisms, predominantly bacteria, which pose potential health risks to spectacle wearers as well as to persons regularly working with worn spectacles, such as opticians [1-3]. ATP measurement represents a rapid and widely used method in hygiene to analyses the organic contamination of surfaces. However, clear correlations with microbial contents on the investigated surfaces are not always provable. Here, we examined the suitability of rapid ATP measurements to assess the efficacy of different cleaning methods for spectacle surfaces. Moreover, we explored the potential correlation with microbial germ counts. To do so, we swab-sampled worn spectacles at our university laboratory, and subsequent ATP bioluminescence assays with a handheld device were used to assess the level of contamination.Six popular cleaning methods were evaluated for their ability to reduce the level of contamination using ten worn spectacles, respectively. Microfiber cloths were the most effective, reducing ATP levels by 93.2%. Alcohol-free wipes followed closely, with a 92.0% decrease. In addition, we investigated the correlation between germ counts and ATP content, which revealed no significant correlation for aerobic cultivation (p = 0.36), but a statistically significant correlation for anaerobic conditions (p = 0.025). Higher CFU values correlated positively with higher ATP levels, suggesting that ATP measurements can rapidly indicate microbial contamination. Our previous studies have shown that spectacles and similar surfaces are indeed colonized by significant shares of (aerotolerant) anaerobic bacteria [2,3]. In summary, ATP measurements are a suitable method to rapidly measure the contamination of spectacles and demonstrate the efficacy of cleaning measures under non-laboratory conditions, such as eyewear stores.[1] Fritz, B. et al. (2018). PLoS ONE 13(11): e0207238. doi.org/10.1371/journal.pone.0207238[2] Fritz B. et al. (2020). Sci Rep. 10(1): 5577. doi.org/10.1038/s41598-020-62186-6.[3] Fritz, B. et al. (2020). J. Clin. Med. 9(5): 1572. doi.org/10.3390/jcm9051572

Abstract: This presentation explores the domain of quality controls in the reprocessing of medical devices, shedding light on current practices, challenges, and avenues for improvement through innovative technological solutions. Medical device reprocessing is integral to ensuring patient safety, reducing healthcare costs, and promoting environmental sustainability. However, it also poses unique challenges related to efficacy, consistency, and regulatory compliance. 
Current quality control practices, including visual inspection and manual cleaning verification, are vital but inherently limited by subjectivity and resource-intensive processes. This highlights the pressing need for innovative technological solutions to augment existing methods and address emerging challenges. 
We explore the design of novel visual inspection systems integrating advanced image recognition technology to automatically identify medical devices, streamlining the inspection workflow, and reducing the risk of human error. Additionally, the design should provide visual assistance to personnel during the quality control process, guiding them through comprehensive checks of sterilizing trays to ensure proper arrangement and integrity of instruments. Practical considerations and challenges associated with the adoption of visual technologies, including integration with existing workflows should also be addressed. 
By leveraging automation, advanced imaging technologies, and data analytics, healthcare facilities can enhance patient safety, streamline operations, and elevate the standard of care in medical device reprocessing. This emphasis on innovation fosters continuous improvement and sets the stage for future advancements in healthcare quality and safety. 

Oral Session
Epidemiology and Antimicrobial Resistance

Appointment

Chair

Topic

Description

Programme

Oral SessionEpidemiology and Antimicrobial Resistance

Appointment

Chair

Topic

Description

Programme

Oral Session
Epidemiology and Antimicrobial Resistance