TY - CONF A1 - Olbrich, Adelina-Elisa T1 - Material vs. Environment: Uncovering microbiological influenced corrosion processes on steel with single cell-ICP-ToF-MS N2 - Microbiologically influenced corrosion (MIC) is a highly unpredictable process dictated by the environment, microorganisms, and the respective electron source. Interaction pathways between cells and the metal surface remain unclear. The development of this novel single cell-inductively coupled plasma-time of flight-mass spectrometry analytical method and a MIC-specific staining procedure facilitate the investigation of steel-MIC interactions. With this it is possible to analyze the multi-elemental fingerprint of individual cells. The detection method revealed elemental selectivity for the corrosive methanogenic archaeal strain Methanobacterium-affiliated IM1. The interface between material and environmental analysis thus receives special attention, e.g., when considering MIC on solid steel. Hence, the possible uptake of individual elements from different steel samples is investigated. Results showed the cells responded at a single-cell level to the different types of supplemented elements and displayed the abilities to interact with chromium, vanadium, titanium, cobalt, and molybdenum from solid metal surfaces. The information obtained will be used in the future to elucidate underlying mechanisms and develop possible material protection concepts, thus combining modern methods of analytical sciences with materials research. T2 - ToFcon 2022 CY - Online meeting DA - 17.11.2022 KW - Single cell KW - Microbiological influenced corrosion MIC KW - Sc-ICP-ToF-MS KW - Method development KW - Ir DNA staining approach KW - Carbon steel corrosion PY - 2022 AN - OPUS4-56355 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Olbrich, Adelina-Elisa T1 - Deciphering microbiological influenced corrosion processes on steel with single cell-ICP-ToF-MS N2 - Microbiologically influenced corrosion (MIC) is a highly unpredictable process dictated by the environment, microorganisms, and the respective electron source. Interaction pathways between cells and the metal surface remain unclear. The development of this novel single cell-inductively coupled plasma-time of flight-mass spectrometry analytical method and a MIC-specific staining procedure facilitate the investigation of steel-MIC interactions. With this it is possible to analyze the multi-elemental fingerprint of individual cells. The detection method revealed elemental selectivity for the corrosive methanogenic archaeal strain Methanobacterium-affiliated IM1. The interface between material and environmental analysis thus receives special attention, e.g., when considering MIC on solid steel. Hence, the possible uptake of individual elements from different steel samples is investigated. Results showed the cells responded at a single-cell level to the different types of supplemented elements and displayed the abilities to interact with chromium, vanadium, titanium, cobalt, and molybdenum from solid metal surfaces. The information obtained will be used in the future to elucidate underlying mechanisms and develop possible material protection concepts, thus combining modern methods of analytical sciences with materials research. References. T2 - Future WiNS CY - Berlin, Germany DA - 07.12.2022 KW - Single cell KW - Microbiological influenced corrosion MIC KW - Sc-ICP-ToF-MS KW - Method development KW - Ir DNA staining approach KW - Carbon steel corrosion PY - 2022 AN - OPUS4-56567 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Razkin Bartolomé, Malen T1 - Bacterial adhesion on fs-laser processed laser-induced periodic surface structures N2 - Bacteria are ubiquitous and colonize all types of surfaces, including those in close proximity to humans, such as skin, food, and everyday objects. This raises the question of whether their presence represents a problem to be mitigated or a potential source of benefit to be harnessed, thereby stimulating scientific inquiry into the role of surface-associated bacteria in diverse domains ranging fromhuman health to industrial biotechnology. Aim: The objective of this project is to explore the impact of modifying surface topography on bacterial adhesion behavior. By manipulating the physical characteristics of the substrate, the attachment and detachment dynamics of bacteria can potentially be modified, leading to novel strategies for controlling bacterial colonization in various applications, such as medical devices. Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) were tested on LIPSS-covered Fused Silica samples. T2 - 2023 Spring Meeting · , 2023 · Strasbourg CY - Strasbourg, France DA - 29.05.2023 KW - LIPSS KW - Biofilm KW - fs-laser processing PY - 2023 AN - OPUS4-58456 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Richter, Anja T1 - Bacterial adhesion on ultrashort laser processed surfaces N2 - Bacterial biofilms are multicellular communities adhering to surfaces and embedded in a self-produced extracellular matrix. Due to physiological adaptations and the protective biofilm matrix itself, biofilm cells show enhanced resistance towards antimicrobial treatment. In medical and industrial settings, biofilms on e.g. for implants or for surfaces in food-processing industry can be a fertile source of bacterial pathogens and are repeatedly associated with persisting, nosocomial and foodborne infections. As extensive usage of antibiotics and biocides can lead to the emergence of resistances, various strategies are currently developed, tested and improved to realize anti-bacterial surface properties through surface functionalization steps avoiding antibiotics. In this study, contact-less and aseptic large-area ultrashort laser scan processing is employed to generate different surface structures in the nanometer- to micrometer-scale on technical materials, i.e. titanium-alloy, steel, and polymer. The processed surfaces were characterized by optical and scanning electron microscopy and subjected to bacterial colonization studies with Escherichia coli test strains. For each material, biofilm results of the fs-laser treated surfaces are compared to that obtained on polished (non-irradiated) surfaces as a reference. Depending on the investigated surfaces, different bacterial adhesion patterns were found, suggesting an influence of geometrical size, shape and cell appendages of the bacteria and – above all – the laser-processed nanostructure of the surface itself. T2 - European Materials Research Society Spring Meeting 2021 CY - Online Meeting DA - 31.05.2021 KW - Bacterial adhesion KW - Biofilm formation KW - Ultrashort laser processing KW - Laser-induced periodic surface structures (LIPSS) PY - 2021 UR - https://www.european-mrs.com/laser-material-processing-fundamental-interactions-innovative-applications-emrs AN - OPUS4-52765 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Richter, Anja T1 - Reducing Escherichia coli adhesion to PET by modulating spatial periods of laser-induced surface nanoripples N2 - Using nanofiber-like cell appendages, secreted proteins and sugars, bacteria can establish initial surface contact followed by irreversible adhesion and the formation of multicellular biofilms. Here, the stabilizing extracellular biofilm matrix together with physiological changes on the single cell level leads to an increased resilience towards harsh environmental conditions, antimicrobials, the host immune response and established cleaning procedures. Persistent microbial adhesion on e.g., medical implants, in water supply networks or food-processing industry is often associated with chronic inflammation, nosocomial and foodborne infections, enhanced biofouling and product contamination. To prevent persistent microbial colonization, antibacterial surface strategies often target the initial steps of biofilm formation and impede adhesion of single cells before a mature biofilm is being formed. While chemical coatings have been widely used, their restricted biocompatibility for eukaryotic cells and attenuated antibacterial-effects due to compound release limit their areas of application and alternative strategies focus on modified surfaces topographies to impede bacterial adhesion. In this work, we used ns-UV laser treatment (wavelength 248 nm and a pulse duration of 20 ns) to generate laser-induced periodic surface structures (LIPSS) with different submicrometric periods ranging from ~210 to ~610 nm on commercial poly(ethylene terephthalate) (PET) foils. Following structurally and chemically analyses, PET samples were subjected to bacterial colonization studies with Escherichia coli TG1, a bacterial test strain with a strong biofilm formation capacity due to the formation of nanofiber-like cell-appendages (pili). Bacterial adhesion tests revealed that E. coli repellence decisively depends on the spatial periods of the LIPSS with the strongest reduction (~91%) in cell adhesion observed for LIPSS periods of 214 nm. Scanning electron microscopy and additional biofilm studies using a pili-deficient E. coli TG1 strain revealed the importance of extracellular appendages in the bacterial repellence observed here, thus, pointing out new antibiotics-free strategies for antibacterial surfaces by impeding nanofiber-mediated bacterial adhesion. T2 - E-MRS Spring Meeting 2022 CY - Online meeting DA - 30.05.2022 KW - Laser-induced periodic surface structures (LIPSS) KW - Bacterial adhesion tests KW - Bacteria repellent surfaces KW - Polymer foils KW - E. coli PY - 2022 AN - OPUS4-54930 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Richter, Anja T1 - Laser-processing – a tool to direct biofilm formation N2 - Using nanofiber-like cell appendages, secreted proteins and sugars, bacteria can establish initial surface contact followed by irreversible adhesion and the formation of multicellular biofilms, often with enhanced resistance towards antimicrobial treatment and established cleaning procedures. On e.g. medical implants, in water supply networks or food-processing industry, biofilms can be a fertile source of bacterial pathogens and are repeatedly associated with persisting, nosocomial and foodborne infections. Nowadays, the emergence of resistances because of extensive usage of antibiotics and biocides in medicine, agriculture and private households have become one of the most important medical challenges with considerable economic consequences. In addition, aggravated biofilm eradication and prolonged cell-surface interaction can lead to increased biodeterioration and undesired modification of industrial and medical surface materials. Various strategies are currently developed, tested, and improved to realize anti-bacterial surface properties through surface functionalization steps avoiding antibiotics. In this study, contact-less and aseptic large-area short or ultrashort laser processing is employed to generate different surface structures in the nanometer- to micrometer-scale on technical materials such as titanium-alloy and polyethylene terephthalate (PET). The laser processed surfaces were subjected to bacterial colonization studies with Escherichia coli test strains and analyzed with reflected-light and epi-fluorescence microscopy. Depending on the investigated surfaces, different bacterial adhesion patterns were found, ranging from bacterial-repellent to bacterial-attractant effects. The results suggest an influence of size, shape and cell appendages of the bacteria and – above all – the laser-processed nanostructure of the surface itself, emphasizing the potential of laser-processing as a versatile tool to control bacterial surface adhesion. T2 - International Biodeterioration & Biodegradation Symposium 2021 CY - Online meeting DA - 06.09.2021 KW - Bacterial adhesion KW - Biofilm formation KW - Laser-induced periodic surface structueres (LIPPS) KW - Laser processing PY - 2021 UR - https://www.ibbs18.org/programme AN - OPUS4-53223 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - THES A1 - Schmidt, Selina T1 - Effects of biocides on processes underlying resistance evolution N2 - Antimicrobial resistance (AMR) is a global health problem. It is well known that antibiotics can drive evolutionary processes that underlie antimicrobial resistance (AMR) evolution and spread in clinical and environmental settings. In contrast, less is known about the effects of antimicrobial substances that are used as biocides (i.e. disinfectants and preservatives) on AMR evolution and spread. Biocides are present in various settings, interacting with diverse microbial communities. Therefore, it is crucial to evaluate their role in the evolution and dissemination of antimicrobial resistance. Biocides occur in a wide range of concentrations in various environmental settings. By examining how the various concentrations affect selection mechanisms, we gain insights into potential developments related to antimicrobial resistance. The aim of this PhD thesis is to investigate the effects of biocides on processes underlying resistance evolution. Specifically, the work focused on key mechanisms for resistance spread, resistance evolution, and the effect of selection pressures on evolved resistance mechanisms. The thesis is structured around three major objectives: (i) to determine the effect of biocides on the evolution of resistance by affecting the rate of occurrence of de novo mutations, (ii) to determine the effect of biocides on the spread of resistance genes by modifying the rate of horizontal gene transfer (HGT) processes, and (iii) to investigate the selective drivers of the emergence of antimicrobial resistance in adaptive laboratory evolution (ALE) experiments. De-novo mutations are spontaneous mutations that occur at a certain rate in microorganisms. The effect of biocides at subinhibitory environmentally relevant concentrations on the mutation rate in Acinetobacer baylyi, Bacillus subtilis and Escherichia coli was assessed with the fluctuation assay. The results showed that biocides affected mutation rates in a species and substance dependent matter. The bisbiguanide chlorhexidine digluconate, the quaternary ammonium compound didecyldimethylammonium chloride, the metal copper, the pyrethroid-insecticide permethrin, and the azole-fungicide propiconazole increase mutation rates in E. coli, whereas no increases were identified for B. subtilis and A. baylyi. Horizontal gene transfer refers to diverse mechanisms that mediate the transfer of mobile genetic elements between microorganisms. This work focused on conjugation and transformation. Conjugation is a process whereby a conjugative plasmid is transferred from a donor cell to a recipient cell. Transformation is a process whereby exogenous donor DNA is taken up into a recipient cell and integrated into the recipient’s’ genome. The effects of subinhibitory environmentally relevant biocide concentrations on the conjugation rate of E. coli and the transformation rate of the naturally competent organisms A. baylyi in were assessed. The results showed that benzalkonium chloride (BAC), chlorhexidine and permethrin increased conjugation in E. coli, while none of the biocides increased transformation rates in A. baylyi. To further understand the molecular mechanisms underlying the effects on mutation and conjugation rates, I investigated the induction of the RpoS-mediated general stress and the RecA-linked SOS response upon biocide exposure. The results show a link between the general stress and the SOS response with increased rates of mutation and conjugation, but not for all biocides. One major approach to study the evolutionary response of bacteria to antimicrobials are ALE experiments with growth at subinhibitory concentrations linked to serial subculturing over many generations. Such experiments have been used to study resistance evolution to antibiotics and biocides. However, previous work showed that adaptation to biocide stress may be mediated by different evolutionary drivers. Here, I investigated the contributions of evolution for increased survival as opposed to improved growth in ALE experiments with E. coli exposed to subinhibitory BAC concentrations. Two distinct evolutionary treatments selecting for survival only or survival and growth led to specific evolutionary adaptations apparent in the phenotypes and genotypes of the evolved populations. Populations growing in the presence of BAC evolved increased fitness in the presence of BAC associated with higher resistance to BAC and cross-resistance to antibiotics, while this was not the case for populations evolving for increased survival only. Genotypic characterization by whole genome sequencing of the evolved populations revealed parallelism in mutated genes among replicate populations and distinct differences across treatments. Treatments selecting for survival and growth showed mutations in stress response related genes (hslO and tufA), while selection for survival led to mutations in genes for metabolic regulation (cyaA) and cellular structure (flagella fliJ). In summary, this thesis shows that biocides affect AMR evolution and emphasizes the importance of understanding of how biocides impact the molecular and evolutionary process that underlie AMR evolution. KW - Biocides KW - Antimicrobial resistances KW - Microbial survival mechanisms PY - 2024 UR - https://nbn-resolving.org/urn:nbn:de:kobv:188-refubium-43383-9 SP - 1 EP - 101 PB - Freie Universität CY - Berlin AN - OPUS4-60678 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schmidt, Selina T1 - Antimicrobial resistance in soil microbes mediated by resistance evolution and horizontal gene transfer (HGT) N2 - Biocides are in contact with soils through direct application and passive leaching from protected materials. Consequently, soil microorganisms are exposed to biocides even though they are not their primary targets. Soil is a large reservoir of microbial diversity and has been hypothesized to be a crucial factor for the evolution and spread of antimicrobial resistance. Currently, there is little knowledge on how biocides used to protect materials affect the evolution and spread of resistance. Thus, our aim is to investigate the risk for the evolution of biocide resistance and cross-resistance to antibiotics. In addition, we aim to elucidate the affect of biocides on the spread of resistance via horizontal gene transfer (HGT). In adaptive laboratory evolution experiments we cultured selected model soil microorganism with representative biocides under selection regimes with increasing and stable biocide concentrations followed by antibiotic and biocide cross-resistance determination. Moreover, we investigate if the selected biocides affect the rates of de novo mutations and HGT of plasmids that carry resistance genes among soil microorganism. Our results show only small increases of biocide resistance during serial transfers under increasing biocide concentrations. One reason for this might be the narrow selective window for biocide resistance due to steep dose-response relationships. Furthermore, our results indicate that a stable low-level biocide regime did not select for high level cross-resistance to antibiotics and other biocides. Moreover, material preservatives affected the rates of HGT via conjugation and the mutation rates at sub-inhibitory concentrations. The results will enable future risk assessment regarding resistance evolution for biocides used as material preservatives. T2 - 4th Evo Eco PhD Meeting CY - Lutherstadt Wittenberg DA - 04.03.2020 KW - Microbiology KW - Biocides KW - Horizontal gene transfer HGT KW - Resistance evolution KW - Antimicrobial resistance PY - 2020 AN - OPUS4-51313 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schmidt, Selina T1 - Biocides Used as Material Preservatives Modify Rates of de novo Mutation and Horizontal Gene Transfer in Bacteria N2 - Biocides used as material preservatives are in contact with various environments during direct application or passive leaching from protected materials. Antimicrobial resistance (AMR) is a global health problem and the environment is an important compartment for the evolution and transmission of AMR. Soil is an environment with a large reservoir of natural microbial communities and antimicrobial resistance (AMR) genes. Those natural microbial communities are frequently exposed to biocides used as material preservatives. Previous studies have shown that antibiotics, metals and pesticides affect the underlying processes of resistance evolution and spread; namely de novo mutagenesis and horizontal gene transfer by conjugation and transformation in microbial communities. However, it is unknown if active substances used in material preservatives are involved in these processes. We show that biocides used as material preservatives affect rates of mutation and conjugation in microorganism in a species- and substance-dependent manner, while rates of transformation are not directly affected. Our data highlights the importance of assessing the contribution of material preservatives on AMR evolution and spread in the environment. T2 - Microbiome Network Meeting CY - Berlin, Germany DA - 20.07.2022 KW - Biocides KW - Horizontal gene transfer KW - Mutation rate PY - 2022 AN - OPUS4-55263 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schmidt, Selina T1 - Bacterial resistance evolution towards disinfectants and antimicrobial surfaces and development of a standardized test N2 - Question Disinfectants and antimicrobial surfaces (AMCs) are important tools to prevent the spread of pathogens and antimicrobial resistant bacteria. However, concerns have been raised about the possibility for the evolution and selection of resistance against disinfectants and AMCs. In turn, resistance against disinfectants and AMCs can be associated to antibiotic resistance due to cross-resistance - a single mechanism conferring resistance to a disinfectant and an antibiotic- and co-resistance - two distinct mechanisms physically linked on e.g. a plasmid. The risk for resistance and cross-resistance during use of biocides (including disinfectants and AMCs) must be evaluated during authorization according to the EU biocidal product regulation. However, to date there is a lack of standardized methods that support risk assessment during the authorization process. Methods We used adaptive laboratory evolution (ALE) experiments which are based on repeated exposure of bacteria to disinfectants or AMCs. The experiments are followed by phenotypic (antimicrobial susceptibility testing) and genotypic (whole genome sequencing) characterization of the evolved strains. The basic idea of these experiments is to expose bacteria to lethal conditions and select for mutants with increased survival. This approach is fundamentally different to other ALE experiments, which commonly select for increased growth at subinhibitory concentrations. However, selection for increased survival represents a selective pressure that more realistically reflects selection under in-use conditions of disinfectants and AMCs. Results First, we studied adaptation of E. coli during repeated disinfection with benzalkonium chloride in a suspension assay. The experiments showed a 2000-fold increase in survival within 5 exposure cycles. The adaptive changes are linked to highly parallel mutations in genes related to lipid A biosynthesis, less negative cell surface charge, reduced growth rate and increased competitive ability in the presence of certain antibiotics. We use the same approach to develop standardizable ALE experiments based upon accepted standards that are used to determine the efficacy of disinfectants (EN 13697) and antimicrobial surfaces (ISO 22196). The results highlight pronounced adaptation of different test strains towards surface disinfection (benzalkonium chloride and isopropanol) and AMCs (copper). Conclusion Bacteria can adapt with increased survival towards lethal stress imposed by disinfectants and AMCs. The adaptive ability of bacteria to disinfectants and AMCs can be determined in a standardized manner. T2 - 74. Jahrestagung der Deutschen Gesellschaft für Hygiene und Mikrobiologie [DGHM] e. V. CY - Berlin, Germany DA - 05.09.2022 KW - Antimicrobial surfaces KW - ISO22196 KW - Antimicrobial resistance KW - Round robin test PY - 2022 AN - OPUS4-56432 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schmidt, Selina T1 - Biocides Used as Material Preservatives Modify Rates of De Novo Mutation And Horizontal Gene Transfer in Bacteria N2 - Antimicrobial resistance (AMR) is an important global health problem. The environment has been regocnized as an improtant compartment for the occurance, evolution and transmission of AMR. Biocides used as material preservatives are in contact with the environment and natural microbial communities through direct application and passive leaching from protected materials. It has been shown that environmental contaminants, such as antibiotics, metals and pesticides, can affect resistance evolution and spread by modifying the underlying processes of de novo mutagenesis, horizontal gene transfer and selection. However, it is unknown if material preservatives are involved in these processes and thereby can also drive AMR in the environment. Here, we investigate the effect of material preservatives on rates of de novo mutation and horizontal gene transfer (HGT) in Escherichia coli and the model soil microorganisms Acinetobacter baylyi and Bacillus subtilis. To this end, we determined the effects of material preservatives on the mutation rates and HGT with the Luria-Delbrück fluctuation assay and a conjugation assay with the broad host-range plasmid pKJK5 and a transformation assay. Our data shows that the quaternary ammonium compound DDAC, copper, the pyrethroid insecticide permethrin and the azole fungicide propiconazole significantly increase mutation rates in E. coli, whereas A. baylyi and B. subtilis are not significantly affected. Moreover, we show that the carbamate IPBC and the insecticide permethrin affect HGT in a concentration dependent manner. Investigations with reporter strains for bacterial stress response pathways show that induction of the general stress response (rpoS) and components of the SOS response (recA) underlie the effects of most biocides on mutation rates and HGT. Taken together, our data is important for assessing the contribution of biocides on AMR evolution and spread in the environment. T2 - World Microbe Forum (ASM, FEMS) CY - Online meeting DA - 20.06.2021 KW - Biocides KW - Horizontal gene transfer KW - Mutation rate PY - 2021 UR - https://www.abstractsonline.com/pp8/#!/9286/presentation/11692 AN - OPUS4-54211 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schmidt, Selina T1 - Effect of biocides on the evolution and spread of resistance N2 - Biocides used as material preservatives are in contact with various environments during direct application or passive leaching from protected materials. Antimicrobial resistance (AMR) is a global health problem and the environment is an important compartment for the evolution and transmission of AMR. Soil is an environment with a large reservoir of natural microbial communities and antimicrobial resistance (AMR) genes. Those natural microbial communities are frequently exposed to biocides used as material preservatives. Previous studies have shown that antibiotics, metals and pesticides affect the underlying processes of resistance evolution and spread; namely de novo mutagenesis and horizontal gene transfer by conjugation and transformation in microbial communities. However, it is unknown if active substances used in material preservatives are involved in these processes. We show that biocides used as material preservatives affect rates of mutation and conjugation in microorganism in a species- and substance-dependent manner, while rates of transformation are not directly affected. Our data highlights the importance of assessing the contribution of material preservatives on AMR evolution and spread in the environment. T2 - Microbiome Network Meeting CY - Berlin, Germany DA - 20.07.2022 KW - Biocides KW - Antimicrobial resistance KW - Horizontal gene transfer KW - Mutation rate PY - 2022 AN - OPUS4-56426 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schmidt, Selina T1 - Effect of biocides on the evolution and spread of resistance N2 - Biocides used as material preservatives are in contact with various environments during direct application or passive leaching from protected materials [1]. Antimicrobial resistance (AMR) is a global health problem and the environment is an important compartment for the evolution and transmission of AMR [2,3]. Soil is an environment with a large reservoir of natural microbial communities and antimicrobial resistance (AMR) genes. Those natural microbial communities are frequently exposed to biocides used as material preservatives. Previous studies have shown that antibiotics, metals and pesticides affect the underlying processes of resistance evolution and spread; namely de novo mutagenesis and horizontal gene transfer by conjugation and transformation in microbial communities. However, it is unknown if active substances used in material preservatives are involved in these processes. We show that biocides used as material preservatives affect rates of mutation and conjugation in microorganism in a species- and substance-dependent manner, while rates of transformation are not directly affected. Our data highlights the importance of assessing the contribution of material preservatives on AMR evolution and spread in the environment. T2 - RokoCon2022 CY - Berlin, Germany DA - 29.09.2022 KW - Biocide KW - Antimicrobial resistance KW - Tolerance KW - Horizontal gene transfer KW - Mutation rate PY - 2022 AN - OPUS4-56431 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Biocide Resistance - Road to Risk Assessment N2 - This presentation details the current status of biocide resistance risk assessment and provides a roadmap for future activities. T2 - OECD, 6th Meeting of the Working Party on Biocides CY - Paris, France DA - 28.09.2022 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Standardization KW - Biocides KW - Risk assessment PY - 2022 AN - OPUS4-56263 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Die Entstehung von antimikrobiellen Resistenzen durch die Verwendung von Bioziden N2 - Dieser Vortrag gibt einen Überblick über die BAM und die Aktivitäten im Bereich Biozidresistenz. T2 - Berliner Hochschule für Technik Studiengang Biotechnologie CY - Berlin, Germany DA - 05.05.2023 KW - Antimikrobielle Resistenz KW - Antmikrobielle Oberflächen KW - Standardisierung PY - 2023 AN - OPUS4-57859 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - How to regulate and assess resistance risks of co-selecting agents during application and in the environment N2 - This talk deals with the question on How to regulate and assess resistance risks of co-selecting agents during application and in the environment. It shows that there the overview of worldwide activities to regulate co-selecting agents is missing. Regulations for product authorization usually consider resistance in target organisms, but there is a risk of emergence of resistance from non-target organisms as well. Moreover, pollution effects on resistance development in the environment are not explicitly covered during product authorization and few risk assessment schemes and methods available. T2 - EDAR7 - Environmental Dimension of Antimicrobial Resistance Conference 2024 CY - Montreal, Canada DA - 26.05.2024 KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides PY - 2024 AN - OPUS4-61542 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Development of a laboratory method to assess resistance development of microorganisms to biocides – An update N2 - This presentation describes the development of a laboratory method to assess resistance development of microorganisms to biocides and antimicrobial surfaces. T2 - The International Biodeterioration Research Group (IBRG) autumn meeting 2022 CY - Online meeting DA - 11.10.2022 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Standardization KW - Biocides KW - Risk assessment PY - 2022 AN - OPUS4-56264 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Development of a laboratory method to assess resistance development of microorganisms to biocides – An update N2 - This presentation describes our efforts at BAM towards the development of a laboratory method to assess resistance development of microorganisms to biocides. T2 - The International Biodeterioration Research Group (IBRG) spring meeting 2023 CY - Online meeting DA - 05.03.2023 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Standardization KW - Biocides PY - 2023 AN - OPUS4-57858 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Development of a laboratory method to assess resistance development of microorganisms to biocides N2 - Bacteria are exposed to biocides through surface disinfection or by antimicrobial surfaces. These stressful environments provide a strong selective pressure for bacteria to adapt. Here, we describe the development of a laboratory method to assess adaption of bacteria by resistance development in response to surface disinfection and antimicrobial surfaces. T2 - OECD, 5th Meeting of the Working Party on Biocides CY - Online meeting DA - 26.05.2021 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Standardization PY - 2021 AN - OPUS4-53163 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Development of a laboratory method to assess resistance development of microorganisms to biocides N2 - Bacteria are exposed to biocides through surface disinfection or by antimicrobial surfaces. These stressful environments provide a strong selective pressure for bacteria to adapt. Here, we describe the development of a laboratory method to assess adaption of bacteria by resistance development in response to surface disinfection and antimicrobial surfaces. T2 - The International Biodeterioration Research Group (IBRG) Spring Meeting 2021 CY - Online meeting DA - 28.06.2021 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Standardization PY - 2021 AN - OPUS4-53164 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Hydro-Touch test - Evaluation of Transfer of Microorganisms to Non-porous Surfaces and their antimicrobial Activity N2 - This presentation describes a new test procedure that allows to determine the transfer of microorganisms to surfaces and to measure the antimicrobial efficacy of those surfaces. The main improvement of the method is that is assesses the effect of the surfaces under semi-dry, realistic conditions by transferring microorganisms via a gelatin pad linked to a stamp of defined weight. Comparing the performance of the established antimicrobial materials with currently used wet test methods and the newly developed method shows reduced antimicrobial activity of those materials under semi-dry conditions. T2 - 64. Sitzung des DIN NA 176-03-06 AA „Chemische Desinfektionsmittel und Antiseptika in der Humanmedizin“ CY - Online meeting DA - 29.09.2025 KW - Antimicrobial surfaces KW - Standardization PY - 2025 AN - OPUS4-64870 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Mechanisms and evolution of resistance to antimicrobial biocides N2 - Antimicrobial resistance (AMR) is a global health problem with the environment being an important compartment for the evolution, selection and transmission of AMR. These processes are impacted by pollution with antibiotics. However, antimicrobial biocides used as disinfectants and material preservatives are major pollutants exceeding the antibiotic market in terms of chemical diversity and mass. The aim of our work is to understand the mechanisms and risks of biocides for resistance and antibiotic cross-resistance evolution in bacteria to optimize their application and safeguard their efficacy. Our work shows that biocides have the potential to affect evolutionary processes towards AMR by increasing the rates of de-novo mutation and conjugation. Importantly, widely used compounds such as chlorhexidine and quaternary ammonium compounds (QACs) affect rates of mutation and conjugation at environmentally relevant concentrations. Furthermore, we show that single-cell phenotypic heterogeneity regarding tolerance (persistence) determines survival against specific biocides including QACs and isopropanol. Mechanistic investigations reveal that known antibiotic persister mechanisms contribute to persister formation to biocides. The evolution of high-level tolerance to different biocides is linked to the initial persister level and the evolution of specific genetically encoded mechanisms related to properties of the cell envelope. Biocide-tolerant strains have a selective advantage in the presence of environmentally-relevant concentrations of antibiotics, which could lead to the stabilization of biocide tolerance in environments where biocides and antibiotics co-occur (e.g. wastewater, animal stables). Taken together, our work shows the importance of assessing the contribution of biocides on evolution and selection of AMR in the environment. T2 - EMBO Symposium on Mechanisms of drug resistance and tolerance in bacteria, fungi, and cancer CY - Heidelberg, Germany DA - 18.03.2025 KW - Antimicrobial surfaces KW - Biocides KW - Antimicrobial resistance PY - 2025 AN - OPUS4-64866 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Resistance evolution towards biocides and antimicrobial surfaces N2 - Biocides, including disinfectants and antimicrobial surfaces (AMCs), are important to prevent the spread of pathogens and antimicrobial resistant bacteria via surfaces. However, concerns have been raised about the evolution and selection of resistance against disinfectants and AMCs. In turn, resistance against disinfectants and AMCs can be associated to antibiotic resistance due to cross-resistance and co-resistance. We need to understand the mechanisms and risks of disinfectants and AMCs for resistance and cross-resistance evolution to optimize their application and safeguard their long-term efficacy. We used adaptive laboratory evolution (ALE) experiments based on repeated exposure of bacteria to disinfectants. Our results show that repeated disinfection of E. coli with benzalkonium chloride in suspension results in a 2000-fold increase in survival within 5 exposure cycles. Adaption is linked to the initial presence of persister cells highly tolerant to benzalkonium chloride. We used the same approach to develop standardizable ALE experiments to determine resistance evolution to AMCs. The results highlight rapid adaptation of E. coli and P. aeruginosa towards copper surfaces. Moreover, there are multiple situations in the clinic or in the environment in which biocides and antibiotics co-occur and in which combination effects can shape their antimicrobial activity or their selective effects. Our work with P. aeruginosa shows prevalent combination effects of biocides and antibiotics, ranging from synergy to antagonism and resulting in the selection for or against antibiotic resistant strains. The combination effects are dependent on the biofilm mode-of-growth, manifesting in apparent differences in the structural arrangement of antibiotic sensitive and resistant strains in biofilms exposed to combinations. Furthermore, biocides affect rates of mutation and horizontal gene transfer, thereby having a potential facilitating effect on resistance evolution. Taken together, our work shows that the role of biocides as potential drivers of resistance evolution and selection deserves further study and regulative action. T2 - Eurobiofilms 2022 CY - Palma, Spain DA - 31.08.2022 KW - Antimicrobial resistance KW - Bacteria KW - Biofilms KW - Biocides KW - Antimicrobial surfaces PY - 2022 AN - OPUS4-55608 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Biocides as drivers for the evolution of antimicrobial resistance N2 - This presentation provides an overview about how biocides drive the evolution of antimicrobial resistance during application and as pollutants in the environment. The presentation shows that biocides can lead to heterogeneous killing, facilitating tolerance evolution. This evolution is related to decreased susceptibility to antibiotics and has potential for co-selection. In contrast, evolved tolerance can limit antibiotic evolvability via epistatic interactions. Moreover, biocides can co-select for antibiotic resistance in wastewater and affect rates of mutation and horizontal gene transfer. Biocides and antibiotics show strong combination effects with consequences for selection of antibiotic resistance. T2 - Novel strategies and considerations in fighting pathogens CY - Tartu, Estonia DA - 16.06.2025 KW - Antimicrobial surfaces KW - Biocides KW - Antimicrobial resistance KW - Standardization PY - 2025 AN - OPUS4-64867 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Biocides as drivers for the Selection and evolution of antimicrobial resistance N2 - Antimicrobial resistance (AMR) is a global health problem with the environment being an important compartment for the evolution, selection and transmission of AMR. These processes are impacted by pollution with antibiotics. However, antimicrobial biocides used as disinfectants and material preservatives are major pollutants exceeding the antibiotic market in terms of chemical diversity and mass. The aim of our work is to understand the mechanisms and risks of biocides for resistance and antibiotic cross-resistance evolution in bacteria to optimize their application and safeguard their efficacy. We use adaptive laboratory evolution experiments, phenotypic characterization, single-cell analysis, whole genome sequencing, and competition experiments to investigate AMR evolution and selection of the model organism E. coli in the presence of biocides. Our work shows that biocides have the potential to affect evolutionary processes towards AMR by increasing the rates of de-novo mutation and conjugation. Importantly, widely used compounds such as chlorhexidine and quaternary ammonium compounds (QACs) affect rates of mutation and conjugation at environmentally relevant concentrations. Furthermore, we show that single-cell phenotypic heterogeneity regarding tolerance (persistence) determines survival against specific biocides including QACs and isopropanol. Mechanistic investigations reveal that known antibiotic persister mechanisms contribute to persister formation to biocides. The evolution of high-level tolerance to different biocides is linked to the initial persister level and the evolution of specific genetically encoded mechanisms related to properties of the cell envelope. Biocide-tolerant strains have a selective advantage in the presence of environmentally-relevant concentrations of antibiotics, which could lead to the stabilization of biocide tolerance in environments where biocides and antibiotics co-occur (e.g. wastewater, animal stables). Taken together, our work shows the importance of assessing the contribution of biocides on evolution and selection of AMR in the environment. T2 - 10th Symposium on Antimicrobial Resistance in Animals and the Environment (ARAE) CY - Berlin, Germany DA - 30.06.2025 KW - Antimicrobial surfaces KW - Biocides KW - Antimicrobial resistance KW - Standardization PY - 2025 AN - OPUS4-64869 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Biocides as drivers for the evolution of antimicrobial resistance N2 - This presentation provides an overview about how biocides drive the evolution of antimicrobial resistance during application and as pollutants in the environment. The presentation shows that biocides can lead to heterogeneous killing, facilitating tolerance evolution. This evolution is related to decreased susceptibility to antibiotics and has potential for co-selection. In contrast, evolved tolerance can limit antibiotic evolvability via epistatic interactions. Moreover, biocides can co-select for antibiotic resistance in wastewater and affect rates of mutation and horizontal gene transfer. Biocides and antibiotics show strong combination effects with consequences for selection of antibiotic resistance. T2 - Vorstellungsvortrag zur Habilitation FU Berlin CY - Berlin, Germany DA - 15.05.2025 KW - Antimicrobial surfaces KW - Biocides KW - Antimicrobial resistance KW - Standardization PY - 2025 AN - OPUS4-64873 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Resistance evolution to antimicrobial biocides N2 - This talk discusses resistance evolution to antimicrobial biocides. It shows (i) that biocides show heterogeneous killing facilitating tolerance evolution, (ii) that serial transfer at subinhibitory concentrations is not appropriate to model evolutionary adaptation to disinfection, (iii) that biocides affect rates of mutation and horizontal gene transfer Biocides, and (iv) that antibiotics show strong combination effects. T2 - Evolutionary Biology meets the Antibiotic Crisis vol. 2 CY - Plön, Germany DA - 24.09.2024 KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides PY - 2024 AN - OPUS4-61545 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Dr. Frank Schreiber and Work on Biocide Resistance at BAM N2 - This talk details the career path of Dr. Frank Schreiber and his work on biocide resistance at BAM. Biocides are antimicrobial products for defined applications (disinfectants, preservatives, pest control). Biocides show heterogeneous killing facilitating resistance/tolerance evolution. Adaptation to biocides has effects on growth and selection. T2 - Vortrag an der Berliner Hochschule für Technik CY - Berlin, Gemany DA - 07.06.2024 KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides PY - 2024 AN - OPUS4-61543 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Biocides as drivers for the evolution and selection of antimicrobial resistance N2 - This presentation provides an overview about how biocides drive the evolution of antimicrobial resistance during application and as pollutants in the environment. The presentation shows that biocides can lead to heterogeneous killing, facilitating tolerance evolution. This evolution is related to decreased susceptibility to antibiotics and has potential for co-selection. In contrast, evolved tolerance can limit antibiotic evolvability via epistatic interactions. Moreover, biocides can co-select for antibiotic resistance in wastewater and affect rates of mutation and horizontal gene transfer. Biocides and antibiotics show strong combination effects with consequences for selection of antibiotic resistance. T2 - 12. Dresdner Wasserseminar ' Wasser und Verunreinigung' CY - Dresden, Germany DA - 26.06.2025 KW - Antimicrobial surfaces KW - Biocides KW - Antimicrobial resistance KW - Standardization PY - 2025 AN - OPUS4-64868 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Beyond Antibiotics – Biocides as Drivers of Environmental AMR N2 - This presentation provides an overview about how biocides drive the evolution of antimicrobial resistance during application and as pollutants in the environment. It also discusses the contribution of biocides for the environmental transmission of AMR. T2 - OneBridge: Making environmental AMR Surveillance Fit for Purpose: Data Integration and the Ecology of Resistance CY - Dresden, Germany DA - 06.10.205 KW - Antimicrobial surfaces KW - Biocides KW - Antimicrobial resistance KW - Standardization PY - 2025 AN - OPUS4-64872 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Resistance evolution towards biocides and antimicrobial surfaces N2 - This presentation describes our work at BAM on resistance evolution towards biocides and antimicrobial surfaces. T2 - Break biofilms workshop CY - Vienna, Austria DA - 16.01.2023 KW - Antimicrobial resistance KW - Bacteria KW - Biofilms KW - Biocides PY - 2023 AN - OPUS4-57857 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Biocide resistance risk assessment N2 - This presentation details the current status of biocide resistance risk assessment. T2 - Risk assessment of biocide and antibiotic resistance CY - Online meeting DA - 09.03.2022 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Standardization KW - Biocides KW - Risk assessment PY - 2022 UR - https://www.gu.se/en/biocide/risk-assessment-of-biocide-and-antibiotic-resistance AN - OPUS4-56235 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Phenotypic and evolutionary responses of bacteria to disinfection N2 - Disinfectants are important to prevent the transmission of pathogens, especially in the face of the current antibiotic resistance crisis. The crisis is further exacerbated by phenotypically tolerant persister subpopulations that can survive transient antibiotic treatment and facilitate resistance evolution. Despite the transient nature of disinfectant application, persistence to disinfectants and its role for the evolution of tolerance and cross-resistance to antibiotics has not been studied. Our work shows that E. coli displays persistence against several widely used disinfectants, including benzalkonium chloride (BAC), didecyldimethylammoniumchlorid (DDAC) and isopropanol. The molecular mechanism of BAC persistence is triggered in stationary phase and affected by several antibiotic persister genes (hipA, tisB, tolC, relA, spoT). Experimental evolution and population dynamic modeling show that repeated failure of disinfection due to persisters rapidly selects for BAC tolerance underpinned by reduced cell surface charge due to mutations in genes related to lipid A acylation (lpxML). Furthermore, evolved BAC tolerance affects the susceptibility to antibiotics, leading to positive selection of disinfectant tolerant strains at environmentally relevant antibiotic concentrations and variations in evolvability of antibiotic resistance due to epistatic effects. These results highlight the need for faithful application of disinfectants to steward their efficacy and the efficacy of antibiotics. A better understanding of the bacterial response to disinfectants is crucial to understand and avert the ongoing antimicrobial resistance crisis. T2 - Gordon Research Conference - Molecular mechanisms of evolution CY - Easton, MA, United States DA - 25.06.2023 KW - Antimicrobial resistance KW - Bacteria KW - Disinfection KW - Biocides PY - 2023 AN - OPUS4-57861 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - BIOCIDE N2 - This presentation gives an overview about the BIOCIDE project performed with the Aquatic Pollutants joint call. T2 - Aquatic Pollutants TransNet workshop CY - Online meeting DA - 09.11.2022 KW - Antimicrobial resistance KW - Bacteria KW - Biofilms KW - Biocides KW - Risk assessment KW - Wastewater PY - 2022 AN - OPUS4-56265 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Resistenzen von Bakterien gegen Biozide – Evolution, Mechanismen und Methoden N2 - Diese Präsentation gibt einen Überblick über die Aktivitäten zum Thema Biozidresistenz an der BAM. T2 - Life Science Nord - Online-Update Hygiene und Infektionsprävention CY - Online meeting DA - 14.06.2022 KW - Antimikrobielle Resistenz KW - Antmikrobielle Oberflächen KW - Standardisierung KW - Biozide KW - Risikobewertung PY - 2022 UR - https://vimeo.com/722198443 AN - OPUS4-56236 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Resistance evolution towards biocides and antimicrobial surfaces N2 - This talk discusses resistance evolution towards biocides and antimicrobial surfaces. It shows (i) that biocides affect rates of mutation and horizontal gene transfer, (ii) that biocides show heterogeneous killing facilitating tolerance evolution, and (iii) that biocides and antibiotics show strong combination effect on growth and selection. T2 - STOP project internal seminar CY - Online meeting DA - 24.04.2024 KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides KW - Antimicrobial surfaces PY - 2024 AN - OPUS4-61546 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Physiological and evolutionary consequences of exposing Pseudomonas aeruginosa to biocide-antibiotic combinations N2 - Antimicrobials can exert specific physiological effects when used in combination that are different from those when applied alone. These effects include physiological effects (i.e. synergy, antagonism and suppression) as well as evolutionary effects on the selection of resistant strains (i.e. cross-resistance and collateral sensitivity). While combination effects have been extensively mapped for antibiotic-antibiotic combinations, the combination effects of antibiotics with antimicrobials used as biocides or antiseptics have not been systematically investigated. Here, we investigated the physiological and evolutionary consequences of combinations of antibiotics (meropenem, gentamicin and ciprofloxacin) and substances used as biocides or antiseptics (octenidine, benzalkonium chloride, cetrimonium bromide, chlorhexidine, povidone-iodine, silver) on growth and selection of Pseudomonas aeruginosa. We find prevalent physiological combination effects with synergy occurring 6 times and antagonism occurring 10 times. The effects are specific to the antibiotic-biocide combination with meropenem showing a tendency for antagonism with biocides (6 of 7), while gentamicin has a tendency for synergy (5 of 7). A particular strong antagonism is apparent for the meropenem-chlorhexidine combination, for which we conducted an in-depth study on the underlying molecular mechanism using RNASeq. Moreover, we find widespread effects of the biocide-antibiotic combinations on selection of P. aeruginosa strains resistant to the antibiotics, including cross-resistance and collateral sensitivity. In conclusion, antibiotics and biocides or antiseptics exert physiological and evolutionary combination effects on the pathogen P. aeruginosa. These effects have consequences for the efficacy of both types of substances and for the selection of antimicrobial resistant strains in clinical applications with combined exposure (e.g. wound care, coated biomaterials). T2 - Antimicrobial Resistance in Biofilms and on Biomaterials CY - Online meeting DA - 10.06.2021 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Biofilms PY - 2021 AN - OPUS4-53162 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Physiological and evolutionary consequences of exposing Pseudomonas aeruginosa to biocide-antibiotic combinations N2 - Antimicrobials can exert specific physiological effects when used in combination that are different from those when applied alone. These effects include physiological effects (i.e. synergy, antagonism and suppression) as well as evolutionary effects on the selection of resistant strains (i.e. cross-resistance and collateral sensitivity). While combination effects have been extensively mapped for antibiotic-antibiotic combinations, the combination effects of antibiotics with antimicrobials used as biocides or antiseptics have not been systematically investigated. Here, we investigated the physiological and evolutionary consequences of combinations of antibiotics (meropenem, gentamicin and ciprofloxacin) and substances used as biocides or antiseptics (octenidine, benzalkonium chloride, cetrimonium bromide, chlorhexidine, povidone-iodine, silver) on growth and selection of Pseudomonas aeruginosa. We find prevalent physiological combination effects with synergy occurring 6 times and antagonism occurring 10 times. The effects are specific to the antibiotic-biocide combination with meropenem showing a tendency for antagonism with biocides (6 of 7), while gentamicin has a tendency for synergy (5 of 7). A particular strong antagonism is apparent for the meropenem-chlorhexidine combination, for which we conducted an in-depth study on the underlying molecular mechanism using RNASeq. Moreover, we find widespread effects of the biocide-antibiotic combinations on selection of P. aeruginosa strains resistant to the antibiotics, including cross-resistance and collateral sensitivity. In conclusion, antibiotics and biocides or antiseptics exert physiological and evolutionary combination effects on the pathogen P. aeruginosa. These effects have consequences for the efficacy of both types of substances and for the selection of antimicrobial resistant strains in clinical applications with combined exposure (e.g. wound care, coated biomaterials). T2 - ASM-FEMS World Microbe Forum CY - Online meeting DA - 20.06.2021 KW - Antimicrobial resistance KW - Antagonism KW - Biofilms PY - 2021 AN - OPUS4-53165 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Risk Assessment of Biocide Resistance N2 - This presentation details approaches for the risk assessment of biocide resistance. Different methods are presented to acquire the necessary data for such risk assessments. T2 - OECD, 7th Meeting of the Working Party on Biocides CY - Leiden, Netherlands DA - 18.09.2023 KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides PY - 2023 AN - OPUS4-59062 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - GEN A1 - Schreiber, Frank T1 - Differential selection for survival and for growth in adaptive laboratory evolution experiments with benzalkonium chloride N2 - Biocides are used to control microorganisms across different applications, but emerging resistance may pose risks for those applications. Resistance to biocides has commonly been studied using adaptive laboratory evolution (ALE) experiments with growth at subinhibitory concentrations linked to serial subculturing. It has been shown recently that E. coli adapts to repeated lethal stress imposed by the biocide benzalkonium chloride (BAC) by increased survival (i.e. tolerance) and not by evolving the ability to grow at increased concentrations (i.e. resistance). Here, we investigate the contributions of evolution for tolerance as opposed to resistance for the outcome of ALE experiments with E. coli exposed to BAC. We find that BAC concentrations close to the half maximal effective concentration (EC50, 4.36 µg mL-1) show initial killing (~40%) before the population resumes growth. This indicates that cells face a two-fold selection pressure: for increased survival and for increased growth. To disentangle the effects of both selection pressures, we conducted two ALE experiments: (i) one with initial killing and continued stress close to the EC50 during growth and (ii) another with initial killing and no stress during growth. Phenotypic characterization of adapted populations showed that growth at higher BAC concentrations was only selected for when BAC was present during growth. Whole genome sequencing revealed distinct differences in mutated genes across treatments. Treatments selecting solely for survival led to mutations in genes for metabolic regulation (cyaA) and cellular structure (flagella fliJ), while treatments selecting for growth and survival led to mutations in genes related to stress response (hslO and tufA). Our results demonstrate that serial subculture ALE experiments with an antimicrobial at sub-inhibitory concentrations can select for increased growth and survival. This finding has implications for the design of ALE experiments to assess resistance risks of antimicrobials in different scenarios such as disinfection, preservation, and environmental pollution. KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides PY - 2024 UR - https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1074740 PB - National Library of Biotechnology Information CY - Bethesda AN - OPUS4-61559 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Bakterielle Resistenzen und Toleranzen gegen Desinfektionsmittel und antimikrobielle Oberflächen N2 - Dieser Vortrag beschreibt die Entstehung von bakteriellen Resistenzen und Toleranzen gegen Desinfektionsmittel und antimikrobielle Oberflächen. T2 - Fachtagung für Krankenhaushygiene der Deutschen Gesellschaft für Krankenhaushygiene CY - Essen, Germany DA - 12.05.2023 KW - Antimikrobielle Resistenz KW - Antmikrobielle Oberflächen KW - Standardisierung KW - Biozide PY - 2023 AN - OPUS4-57860 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Biocides As Drivers For Antimicrobial Resistance Evolution In The Environment N2 - Antimicrobial resistance (AMR) is a global health problem with the environment being an important compartment for the evolution, selection and transmission of AMR. These processes are impacted by pollution with antibiotics. However, biocides used as disinfectants and material preservatives are major pollutants by far excceding the market for antibiotics in terms of mass. Our work shows that biocides have the potential to affect evolutionary processes towards AMR by increasing the rates of de-novo mutation and conjugation. These effects depend on the species and biocidal substance. Importantly, chlorhexidine and quaternary ammonium compounds (QACs) affect rates of mutation and conjugation at environmentally relevant concentrations in E. coli. Moreover, our results show a connection between the RpoS-mediated general stress and the RecA-linked SOS response with increased rates of mutation and conjugation, but not for all biocides. Furthermore, our work highlights the potential of biocides to contribute to selection and transmission of AMR. We show that the application of biocides, especially QAC disinfectants, leads to the rapid evolution of tolerance (i.e. increased survival) in adaptive laboratory evolution (ALE) experiments. The evolved tolerant strains have a selective advantage in the presence of environmentally-relevant concentrations of antibiotics, which could lead to the stabilization of biocide tolerance in environments where biocides and antibiotics co-occur (e.g. wastewater, animal stables). ALE experiments with biocide tolerant strains indicate a decreased evolvability of resistance to antibiotics. Taken together, our work shows the importance of assessing the contribution of biocides on evolution, selection and transmission of AMR in the environment. T2 - 6th Environmental Dimension of Antibiotic Resistance (EDAR6) CY - Gothenburg, Sweden DA - 22.09.2022 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Biofilms KW - Biocides KW - Risk assessment PY - 2022 AN - OPUS4-56262 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Resistance evolution towards biocides and antimicrobial surfaces N2 - This talk discusses the work at BAM concerning Resistance evolution towards biocides and antimicrobial surfaces. It shows (i) that biocides affect rates of mutation and horizontal gene transfer, (ii) that biocides show heterogeneous killing facilitating tolerance evolution, and (iii) that biocides and antibiotics show strong combination effect on growth and selection. T2 - Exchange seminar with Nottingham Trent University CY - Berlin, Germany DA - 03.09.2024 KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides KW - Antimicrobial surfaces PY - 2024 AN - OPUS4-61548 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - GEN A1 - Schreiber, Frank T1 - Differential selection for survival and for growth in adaptive laboratory evolution experiments with benzalkonium chloride N2 - Biocides are used to control microorganisms across different applications, but emerging resistance may pose risks for those applications. Resistance to biocides has commonly been studied using adaptive laboratory evolution (ALE) experiments with growth at subinhibitory concentrations linked to serial subculturing. It has been shown recently that E. coli adapts to repeated lethal stress imposed by the biocide benzalkonium chloride (BAC) by increased survival (i.e. tolerance) and not by evolving the ability to grow at increased concentrations (i.e. resistance). Here, we investigate the contributions of evolution for tolerance as opposed to resistance for the outcome of ALE experiments with E. coli exposed to BAC. We find that BAC concentrations close to the half maximal effective concentration (EC50, 4.36 µg mL-1) show initial killing (~40%) before the population resumes growth. This indicates that cells face a two-fold selection pressure: for increased survival and for increased growth. To disentangle the effects of both selection pressures, we conducted two ALE experiments: (i) one with initial killing and continued stress close to the EC50 during growth and (ii) another with initial killing and no stress during growth. Phenotypic characterization of adapted populations showed that growth at higher BAC concentrations was only selected for when BAC was present during growth. Whole genome sequencing revealed distinct differences in mutated genes across treatments. Treatments selecting solely for survival led to mutations in genes for metabolic regulation (cyaA) and cellular structure (flagella fliJ), while treatments selecting for growth and survival led to mutations in genes related to stress response (hslO and tufA). Our results demonstrate that serial subculture ALE experiments with an antimicrobial at sub-inhibitory concentrations can select for increased growth and survival. This finding has implications for the design of ALE experiments to assess resistance risks of antimicrobials in different scenarios such as disinfection, preservation, and environmental pollution. KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides PY - 2024 DO - https://doi.org/10.5061/dryad.2jm63xszx PB - Dryad AN - OPUS4-61558 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank T1 - Selection of resistance by antimicrobials used in coatings N2 - Antimicrobial surfaces have broad use in multiple settings including touch surfaces in hospitals, implanted devices, or consumer products. Their aim is to support existing hygiene procedures, and to help combat the increasing threat of antimicrobial resistance. However, concerns have been raised over the potential selection pressure exerted by such surfaces, which might drive the evolution and spread of antimicrobial resistance. In my presentation, I will highlight the risks and knowledge gaps associated with resistance on antimicrobial surfaces by different processes including evolution by de novo mutations and horizontal gene transfer, and species sorting of inherently resistant bacteria dispersed onto antimicrobial surfaces. The latter process has the potential to select for antibiotic resistance via cross-resistance between traits that confer resistance to both the antimicrobial surface coating and antibiotics. Conditions in which antibiotics and antimicrobial coatings are present simultaneously (e.g. implants) will lead to more complex interactions that can either result in the selection for or against antibiotic resistance. We mapped these interactions between several antimicrobials and antibiotics on growth and selection of Pseudomonas aeruginosa. We find prevalent physiological (i.e. synergy and antagonism) and evolutionary (i.e. cross-resistance and collateral sensitivity) combination effects. Understanding these interactions opens the door to tailor therapeutic interventions to select against resistance. In additions, we need new methods and translational studies that investigate resistance development to antimicrobial surfaces under realistic conditions. Therefore, I will present recent developments in our lab on the development of such a method based on existing efficacy standards. T2 - 2021 Fall Meeting of the European Materials Research Society (E-MRS) CY - Online meeting DA - 20.09.2021 KW - Antimicrobial resistance KW - Antimicrobial coating KW - Biofilms KW - Biocides PY - 2021 AN - OPUS4-53645 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - GEN A1 - Schreiber, Frank A1 - Kanaris, Orestis A1 - Nordholt, Niclas T1 - Consequences of benzalkonium chloride tolerance for selection dynamics and de novode novo resistance evolution driven by antibiotics N2 - Biocides are used in large amounts in industrial, medical, and domestic settings. Benzalkonium chloride (BAC) is a commonly used biocide, for which previous research revealed that Escherichia coli can rapidly adapt to tolerate BAC-disinfection, with consequences for antibiotic susceptibility. However, the consequences of BAC- tolerance for selection dynamics and resistance evolution to antibiotics remain unknown. Here, we investigated the effect of BAC -tolerance in E. coli on its response upon challenge with different antibiotics. Competition assays showed that subinhibitory concentrations of ciprofloxacin—but not ampicillin, colistin and gentamicin—select for the BAC-tolerant strain over the BAC-sensitive ancestor at a minimal selective concentration of 0.0013–0.0022 µg∙mL−1. In contrast, the BAC-sensitive ancestor was more likely to evolve resistance to ciprofloxacin, colistin and gentamicin than the BAC-tolerant strain when adapted to higher concentrations of antibiotics in a serial transfer laboratory evolution experiment. The observed difference in the evolvability of resistance to ciprofloxacin was partly explained by an epistatic interaction between the mutations conferring BAC -tolerance and a knockout mutation in ompF encoding for the outer membrane porin F. Taken together, these findings suggest that BAC -tolerance can be stabilized in environments containing low concentrations of ciprofloxacin, while it also constrains evolutionary pathways towards antibiotic resistance. KW - Antimicrobial surfaces KW - Biocides KW - Antimicrobial resistance KW - Antibiotics PY - 2025 UR - https://www.ncbi.nlm.nih.gov/bioproject/?term=PRJNA1282584 PB - National Library of Medicine CY - Bethesda AN - OPUS4-65221 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schreiber, Frank A1 - Nordholt, Niclas T1 - Persistence against benzalkonium chloride promotes rapid evolution of tolerance under periodic disinfection N2 - Biocides used as disinfectants are important to prevent the transmission of pathogens, especially during the current antibiotic resistance crisis. This crisis is exacerbated by phenotypically tolerant persister subpopulations which can survive transient antibiotic treatment and facilitate resistance evolution. Despite the transient nature of disinfection, knowledge concerning persistence to disinfectants and its link to resistance evolution is currently lacking. Here, we show that E. coli displays persistence against a widely used disinfectant benzalkonium chloride (BAC). Periodic, persister-mediated failure of disinfection rapidly selects for BAC tolerance. BAC tolerance is associated with reduced cell surface charge and mutations in the novel tolerance locus lpxM. Moreover, the fitness cost incurred by BAC tolerance turned into a fitness benefit in the presence of antibiotics, suggesting a selective advantage of BAC-tolerant mutants in antibiotic environments. Our findings provide a mechanistic underpinning for the faithful application of disinfectants to prevent multi-drug-resistance evolution and to steward the efficacy of biocides and antibiotics. T2 - New Approaches and Concepts in Microbiology CY - Online meeting DA - 07.07.2021 KW - Persistence KW - Biocides KW - Evolution KW - Cross-resistance KW - Biocide tolerance PY - 2021 AN - OPUS4-53168 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - INPR A1 - Schreiber, Frank A1 - Nordholt, Niclas A1 - Lewerenz, Dominique T1 - Time-kill kinetics reveal heterogeneous tolerance to disinfectants N2 - Disinfection is an important strategy to limit the spread of infections. Failure of disinfection may facilitate evolution of resistance against disinfectants and antibiotics through the processes of cross-resistance and co-resistance. The best possible outcome of disinfection minimizes the number of surviving bacteria and the chance for resistance evolution. Resistance describes the ability to grow in previously inhibitory concentrations of an antimicrobial, whereas tolerance is associated with enhanced survival of lethal doses. Individual bacteria from the same population can display considerable heterogeneity in their ability to survive treatment (i.e. tolerance) with antimicrobials, which can result in unexpected treatment failure. Here, we investigated how phenotypic heterogeneity affects the ability of E. coli to survive treatment with six different substances commonly used as active substances in disinfectants, preservatives and antiseptics. A mathematical model which assumes that phenotypic heterogeneity underlies the observed disinfection kinetics was used to infer whether time-kill kinetics were caused by a tolerant subpopulation. The analysis identified bimodal kill kinetics for benzalkonium chloride (BAC), didecyldimethylammonium chloride (DDAC), and isopropanol (Iso). In contrast, kill kinetics by chlorhexidine (CHX), glutaraldehyde (GTA), and hydrogen peroxide (H2O2) were best explained by unimodal kill kinetics underpinned by a broad distribution of tolerance times for CHX as opposed to a narrow distribution of tolerance times for GTA and H2O2. These findings have implications for the risk of disinfection failure, with potential consequences for the evolution of antimicrobial resistance and tolerance. KW - Antimicrobial resistance KW - Bacteria KW - Standardization KW - Biocides PY - 2022 UR - https://nbn-resolving.org/urn:nbn:de:kobv:b43-615572 DO - https://doi.org/https://doi.org/10.1101/2022.06.22.497202 SN - 2692-8205 SP - 1 EP - 20 PB - Cold Spring Harbor Laboratory CY - Cold Spring Harbor, NY AN - OPUS4-61557 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schulz, Anne-Christine T1 - Co-selection for biocide and antibiotic resistance in microbial wastewater communities N2 - Biocides are used for a wide range of purposes, including disinfectants or preservatives. They play a major role in the prevention of microbial infections in healthcare and animal husbandry. The use of biocides often leads to the discharge of active biocidal substances into wastewater streams, causing the exposure of wastewater microbial communities to subinhibitory concentrations. In turn, it is known that wastewater treatment plants (WWTP) are hotspots for antibiotic resistant bacteria. Since similar mechanisms confer resistance to biocides and antibiotics, exposure to biocides can result in co-selection of antibiotic resistant bacteria in WWTP Here, we want to investigate co-selection processes of antibiotic resistance in natural WWTP microbial communities upon biocide exposure. Microbial communities were sampled at the WWTP Ruhleben in Berlin and characterized regarding their susceptibility against different clinically relevant antibiotics. To investigate the link between biocide exposure and antibiotic resistance, changes in the susceptibility level after exposure to environmentally relevant concentrations of the commonly used biocide didecyldimethylammonium chloride (DDAC) will be determined by enumerating resistant and non-resistant E. coli on selective plates with and without antibiotics and DDAC. In case of antibiotics, clinical breakpoint concentrations according to EUCAST will be used to discriminate between susceptible and resistant strains. In case of DDAC (and biocides in general), clinical breakpoints do not exist. Therefore, we determined a cut-off concentration at which the majority of naturally-occurring E. coli strains cannot grow anymore based on (I) the MIC (minimal inhibitory concentration) distribution, and (II) by plating wastewater communities onto selective indicator agar plates loaded with increasing DDAC concentration. Additionally, antibiotic cross-resistance will be determined by spotting single colonies, isolated from DDAC-selective plates onto antibiotic plates. The results of our experiments will help to determine selective concentrations and to estimate the risk of antibiotic co-selection and cross-resistance in microbial WWTP communities upon biocide exposure. T2 - Annual Conference 2023 of the Association for General and Applied Microbiology CY - Göttingen, Germany DA - 10.09.2023 KW - Antibiotic resisitance KW - Biocide KW - Wastewater KW - Risk assesment PY - 2023 AN - OPUS4-58510 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Schulz, Anne-Christine T1 - Assessing co-selection of biocide and antibiotic resistance in wastewater microbial communities N2 - Biocides are used for a wide range of purposes, including disinfectants or preservatives. Biocides play a major role in the prevention of microbial infections in healthcare and animal husbandry. The use of biocides often leads to the discharge of active biocidal substances into wastewater streams, causing the exposure of wastewater microbial communities to subinhibitory concentrations. In turn, it is known that wastewater treatment plants (WWTP) are hotspots for antibiotic resistant bacteria. Since similar mechanisms confer resistance to biocides and antibiotics, exposure to biocides can result in co-selection of antibiotic resistant bacteria in WWTP due to biocides. Here, we investigate the magnitude and the drivers of co-selection of antibiotic resistance in natural wastewater microbial communities upon biocide exposure. Microbial communities will be sampled at the WWTP Ruhleben in Berlin and characterized regarding their biocide and antibiotic resistance. Changes in the resistance level after exposure to different biocides will be determined by enumerating resistant and non-resistant E. coli and heterotrophic bacteria on selective plates with and without several biocides and antibiotics. Moreover, we are establishing a synthetic community comprising about 100 environmental E. coli isolates each with different antimicrobial resistance traits. Each isolate will be tagged with a unique DNA-barcode. All isolates will be pooled and exposed to different biocides at various concentrations. The barcode labeling enables us to determine the abundance of each isolate at the beginning and end of the experiment by transposon-tag sequencing. The project results will inform risk assessment of the effects of biocidal residues on antimicrobial resistance selection in WWTP. The project is part of the BIOCIDE consortium funded within the call on Aquatic pollutants by JPI-AMR, JPI-OCEANS and JPI-WATER. T2 - 6th International Symposium onn the Environmental Dimension of Antibiotic Resistance - EDAR 6 CY - Gothenburg, Sweden DA - 22.09.2022 KW - Antibiotic resisitance KW - Biocide KW - Wastewater KW - Risk assesment PY - 2022 AN - OPUS4-56796 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER -