The Physics of Antimicrobial Resistance

Lead Research Organisation: University of Sheffield
Department Name: Physics and Astronomy

Abstract

The development by bacteria of resistance to antibiotics (antimicrobial resistance, AMR) is a global challenge that threatens to undermine many of the advances of modern medicine, with consequential massive human and financial costs. AMR is a multi-faceted problem in which processes occurring over many different length and timescales interact, leading to the emergence of resistant bacteria. To obtain a predictive understanding of this complexity we will take an interdisciplinary approach, bringing together quantitative experimental and mathematical physics with cutting-edge microbiology, biochemistry and infectious disease biology. Bacteria become resistant through genetic mutation and gene acquisition which inevitably leads to physiological changes, including the obvious sustained growth when under antibiotic stress. By better understanding the physical nature of these changes we aim to reveal exploitable fitness costs associated with AMR, i.e. ways in which the bacteria become more vulnerable as the price they pay for becoming resistant to particular antibiotics.

Our programme will focus on resistance mechanisms to cell wall targeting antibiotics. Bacteria have a cell wall that keeps them alive which is made from peptidoglycan, a material not produced by humans. Antibiotics such as penicillin which target the synthesis of peptidoglycan are clinically critically important. Bacterial strains resistant to these antibiotics, such as methicillin resistant S. aureus (the "hospital superbug" MRSA) and carbapenem-resistant enterobacteriaceae (CRE) are recognised as global threats. Concentrating our efforts on these WHO priority organisms provides a direct translational route for the programme.

To provide breakthroughs in understanding we will take a multi-pronged approach. We will combine cutting edge atomic force microscopy, development of new instrumentation as required, state-of-the-art biochemistry and mechanical modelling to find how the cell wall differs between bacteria sensitive and resistant to particular antibiotics. The relationship between chemistry, molecular organisation, and physical properties, is a problem at the heart of materials physics, and here, by correlating quantitative experiments with molecular modelling we will provide a predictive understanding of the cell wall and how it is changed by resistance. Secondly, we will concentrate on how AMR alters bacterial physiology. For example, MRSA has acquired a new enzyme for cell wall synthesis, circumventing the need for the native, antibiotic sensitive target of beta-lactam antibiotics (such as penicillin). We have shown that this enzyme alone is not enough to be resistant; there needs to be additional changes to the transcription machinery (RNA polymerase). Using single molecule and statistical physics approaches that we will adapt and advance for this problem, coupled with molecular biology and biochemistry, we will gain an understanding of these interconnected webs of interaction that drive resistance evolution and characterise AMR organisms. Thirdly, we will explore how AMR impacts bacterial fitness under different conditions, using a combination of state-of-the-art microfluidics based in vitro experiments with in vivo experiments to ensure relevance to the real conditions in a living host. Hence we will find conditions under which AMR organisms are vulnerable to targeted treatments.

To reach our ambitious goals we have brought together a unique team with experts in atomic force microscopy, single molecule biophysics, microfluidics and theoretical physics, in the microbiology of Gram negative and Gram positive bacteria, and in the biochemistry of transcription. Taking an integrated approach, the project will provide a new understanding of AMR with direct clinical relevance.

Planned Impact

The impacts of this Programme our exceptionally wide ranging. AMR is a critically important global health challenge and our Programme aims to develop new insights and understanding that will directly impact human health nationally and internationally. The pharmaceutical industry is an important sector of the UK economy, and our project has the potential to have positive impact in this area also. To reach our ambitious scientific goals we will develop new techniques and approaches which will be of wider application and hence commercial value, and we will aim to exploit these possibilities through the project. We will train PDRAs in cutting edge interdisciplinary science which will have long term impact on their careers, and publicise our work and approach to the public to help inspire the next generation of researchers. Below we provide more detail on these specific areas.

Health in our society: Antibiotics are a cornerstone of modern medicine, allowing treatment of infections but also enabling many of the other treatments that we take for granted, such as chemotherapy, elective surgery, etc., as antibiotics prevent complications. AMR poses an extraordinary threat the potential impact of which is hard to over-state. Our Programme, by taking an interdisciplinary approach that brings together the predictive capabilities of theoretical physics with cutting edge biology and biophysics, provides an opportunity to make a real difference to how we approach antibiotic stewardship and how we treat AMR.

The pharmaceutical industry: The Programme team are exceptionally well connected across the UK and wider pharmaceutical industry. SF was founding director of a S. aureus vaccine company, Absynth Biologics Ltd. and is well placed to further exploit opportunities as they arise. WV and NZ work closely with Demuris Ltd, a Newcastle based drug discovery company, on potential new antibiotic targets. Our work may also reveal opportunities to re-purpose existing drugs to exploit AMR induced fitness changes. Hence our Programme will contribute to the re-invigoration of the UK pharmaceutical industry.

New techniques and approaches: Our Programme will use and where necessary develop cutting edge techniques for studying bacteria across scales from individual molecules to host organisms. Our target is AMR, but these approaches will doubtless have wider applicability, and we will ensure that as wide as possible an audience is reached. JH is highly experienced in commercialising opportunities within AFM, having founded the successful silicon surface inspection company Infinitesima Ltd., and we are well placed to maximise impact in this area.

Our Programme will be world leading example of interdisciplinary research, spanning multiple length and timescales and using an exceptionally wide range of complementary approaches. The PDRAs involved in the project will gain a unique set of skills which will make them highly employable and help to fulfil the UK requirement for scientists capable of applying quantitative approaches to biological systems and data. We will also showcase the Physics of Life Programme as an example of the best that interdisciplinarity can bring to science. We will impress on the next generation of potential researchers, teachers and industrial leaders, how they need not be constrained by traditional discipline boundaries but can be driven by the desire to understand how life works using whatever tools and approaches are necessary.

Publications

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Salamaga B (2021) Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics. in Proceedings of the National Academy of Sciences of the United States of America

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Stevenson-Jones F (2020) Ribosome reactivates transcription by physically pushing RNA polymerase out of transcription arrest. in Proceedings of the National Academy of Sciences of the United States of America

 
Description We have found that the surface of the cell wall of Gram positive bacteria that surrounds and protects it from its environment has a surprising porous texture a bit like a sponge, with holes penetrating almost all the way through to the membrane that it supports. In contrast the internal surface of the cell wall has a dense mesh structure with very tiny pores, just a few nanometres across. When treated with antibiotics the cell can no longer able to make new cell wall, but continues to re-model the cell wall that already exists. This leads to the internal pores becoming larger and joining up with the external holes, damaging the structure of the wall and eventually leading to death.

We have also found that some of the techniques we have developed can be used for studying fungal cell walls, about which relatively little is understood. This is now being taken forward by two PhD students in collaboration with Syngenta.

We have determined the key components of resistance to methicillin in MRSA and this work is currently under consideration for publication.

We have determined the processes that occur during the antibiotic (mecillinam) induced death of E. coli at a molecular level within the cell wall. This work is currently being prepared for publication.

We have unravelled the role of a suite of processes in the resistance of mecillinam resistant E. coli. This work is currently being prepared for publication.
Exploitation Route Understanding how antibiotics kill bacteria can help us to develop new therapies to combat antimicrobial resistance. The approaches developed for imaging cell walls are being taken forward by ourselves and others, including for the exploration of fungal cell walls. The analysis approaches of AFM force data have been taken forward by ourselves in collaboration with Infinitesima Ltd for the exploration of deep trenches in semiconductor devices, leading to patent protection of this approach.
Sectors Agriculture

Food and Drink

Healthcare

 
Description The data analysis approaches developed as part of the project contributed to an improved understanding of tip-sample interactions in AFM and subsequently led to a patent application. This is being utilised by Infinitesima Ltd develop new markets for the microscopy tools.
First Year Of Impact 2022
Sector Digital/Communication/Information Technologies (including Software)
Impact Types Economic

 
Description A Matter Of Life Or Death - An Integrated Understanding Of MRSA
Amount £5,990,700 (GBP)
Funding ID 227233/Z/23/Z 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 01/2024 
End 12/2029
 
Description Bacterial Cell Envelope Biogenesis
Amount £476,610 (GBP)
Funding ID BB/W005557/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 02/2022 
End 01/2025
 
Description Regulation of Autolysins
Amount £427,119 (GBP)
Funding ID BB/W013630/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 06/2022 
End 06/2025
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Figure 1 
Description Dataset used to create Figure 1 in paper accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Figure 1 
Description Dataset used to create Figure 1 in paper accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Figure 2 
Description Dataset used to create Figure 2 in paper accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Figure 2 
Description Dataset used to create Figure 2 in paper accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Figure 3 
Description Dataset used to create Figure 3 in paper accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Figure 3 
Description Dataset used to create Figure 3 in paper accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Figure 4 
Description Dataset used to create Figure 4 in paper accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Figure 4 
Description Dataset used to create Figure 4 in paper accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Supplementary Information 
Description Dataset used to create supplementary information for article accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. Animal work was completed in accordance to UK law in the Animals (Scientific Procedures) Act 1986, under Project Licence P3BFD6DB9 for murine experiments, with approval of the Sheffield Ethical Review Committee. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics: Supplementary Information 
Description Dataset used to create supplementary information for article accepted for publication in PNAS: 'Demonstration of the role of cell wall homeostasis in Staphylococcus aureus growth and the action of bactericidal antibiotics'. Animal work was completed in accordance to UK law in the Animals (Scientific Procedures) Act 1986, under Project Licence P3BFD6DB9 for murine experiments, with approval of the Sheffield Ethical Review Committee. 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Demonstration_of_the_role_of_cell_wall_homeostasis_in_S...
 
Title Evolving MRSA: High-level ß-lactam resistance in Staphylococcus aureus is associated with RNA Polymerase alterations and fine tuning of gene expression 
Description This is the collection of data supporting all the findings in our manuscript, details of which will be available on publication. This includes raw data used to analyse and visualise the outcomes. Raw data associated with each figure are provided in Excel files with corresponding README text on the first tab. 
Type Of Material Database/Collection of data 
Year Produced 2020 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Evolving_MRSA_High-level_-lactam_resistance_in_Staphylo...
 
Title Evolving MRSA: High-level ß-lactam resistance in Staphylococcus aureus is associated with RNA Polymerase alterations and fine tuning of gene expression 
Description This is the collection of data supporting all the findings in our manuscript, details of which will be available on publication. This includes raw data used to analyse and visualise the outcomes. Raw data associated with each figure are provided in Excel files with corresponding README text on the first tab. 
Type Of Material Database/Collection of data 
Year Produced 2020 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Evolving_MRSA_High-level_-lactam_resistance_in_Staphylo...
 
Title Penicillin-Binding Protein 1 (PBP1) of Staphylococcus aureus Has Multiple Essential Functions in Cell Division: Figures 1-5 
Description Dataset used to create supplementary information for article accepted for publication in mBio 'Penicillin-Binding Protein 1 (PBP1) of Staphylococcus aureus Has Multiple Essential Functions in Cell Division'. 
Type Of Material Database/Collection of data 
Year Produced 2022 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Penicillin-Binding_Protein_1_PBP1_of_Staphylococcus_aur...
 
Title Penicillin-Binding Protein 1 (PBP1) of Staphylococcus aureus Has Multiple Essential Functions in Cell Division: Figures 1-5 
Description Dataset used to create supplementary information for article accepted for publication in mBio 'Penicillin-Binding Protein 1 (PBP1) of Staphylococcus aureus Has Multiple Essential Functions in Cell Division'. 
Type Of Material Database/Collection of data 
Year Produced 2022 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Penicillin-Binding_Protein_1_PBP1_of_Staphylococcus_aur...
 
Title Penicillin-Binding Protein 1 (PBP1) of Staphylococcus aureus Has Multiple Essential Functions in Cell Division: Supplementary Figures S1-S8 
Description Dataset used to create supplementary information for article accepted for publication in mBio 'Penicillin-Binding Protein 1 (PBP1) of Staphylococcus aureus Has Multiple Essential Functions in Cell Division'. 
Type Of Material Database/Collection of data 
Year Produced 2022 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Penicillin-Binding_Protein_1_PBP1_of_Staphylococcus_aur...
 
Title Penicillin-Binding Protein 1 (PBP1) of Staphylococcus aureus Has Multiple Essential Functions in Cell Division: Supplementary Figures S1-S8 
Description Dataset used to create supplementary information for article accepted for publication in mBio 'Penicillin-Binding Protein 1 (PBP1) of Staphylococcus aureus Has Multiple Essential Functions in Cell Division'. 
Type Of Material Database/Collection of data 
Year Produced 2022 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Penicillin-Binding_Protein_1_PBP1_of_Staphylococcus_aur...
 
Title Research data in support of "A synthetic signalling network imitating the action of immune cells in response to bacterial metabolism" 
Description This repository contains raw data in support of the publication 10.1002/adma.202301562. Please refer to the published paper, available open access, for full information and context. Figure 2 data - Panel b: Confocal image of core-shell particle - Panel d: Absorbance data to probe pH responsiveness of the DNA constructs - Panel c: CD data to probe pH responsiveness of the DNA constructs - Panel g: Representative epifluorescence and bright field images and bright field videos to demonstrate pH-induced particle aggregation Figure 3 data - Panel b: pH and OD data vs E. coli growth time in various glucose concentrations - Panels d, e; Representative epifluorescence images and bright field videos to demonstrate E. coli induced particle aggregation and bacteria trapping - Panel f: OD data vs E. coli growth time Figure 4 data - Panel b: OD data vs E. coli growth time showing effect of netosis-like synthetic pathway - Panel e: Representative bright field and epifluorescence images showing effect of netosis-like synthetic pathway Supplementary Figure 1: Agarose gel demonstrating DNA construct assembly Supplementary Figure 2: DLS data showing pH responsiveness of DNA nanostructures Supplementary Figure 3: UV melting curve data for DNA constructs Supplementary Figure 4: DLS data underpinning pH response curve Supplementary Figure 5: Representative data provided in Figure 2, Panel g folder Supplementary Figure 6: Representative microscopy videos used to generate hydrodynamic radius data with DDM Supplementary Figure 7: OD data vs E. coli growth time at various glucose concentrations Supplementary Figure 8: FITC dextran fluorescence intensity data (used to determine pH) vs E. coli growth time at various glucose concentrations Supplementary Figure 9: Calibration data (FITC dextran fluorescence vs pH) used in Figure 3f and Supplementary Figures 8 and 12 Supplementary Figure 10: FITC dextran fluorescence intensity data (used to determine pH in Figure 3f) vs E. coli growth time at various glucose concentrations Supplementary Figure 11: Representative data provided in Figure 3, Panel d, e folder Supplementary Figure 12: FITC dextran fluorescence intensity data (used to determine pH) vs E. coli growth time Supplementary Figure 13: Calibration data (Fluorescein dT fluorescence vs pH) for fluorescent pH probe linked to DNA particles Supplementary Figure 14: Fluorescent data for pH-induced de-quenching on DNA particles Supplementary Figure 15: OD vs E. coli growth time at various antibiotic concentrations Supplementary Figure 16: Representative data provided in Figure 4, Panel c, folder 
Type Of Material Database/Collection of data 
Year Produced 2023 
Provided To Others? Yes  
URL https://www.repository.cam.ac.uk/handle/1810/353635
 
Title Staphylococcus aureus cell wall structure and dynamics during host-pathogen interaction 
Description This is the raw data supporting the findings (both main text and supplementary) for our manuscript "Staphylococcus aureus cell wall structure and dynamics during host-pathogen interaction". Each excel file contains the raw data for each figure. Murine work was carried out according to UK law in the Animals (Scientific Procedures) Act 1986, under Project License P3BFD6DB9 (Staphylococcus aureus and other pathogens, pathogenesis to therapy, University of Sheffield Review Board). 
Type Of Material Database/Collection of data 
Year Produced 2021 
Provided To Others? Yes  
URL https://figshare.shef.ac.uk/articles/dataset/Staphylococcus_aureus_cell_wall_structure_and_dynamics_...
 
Title COMPOUNDS FOR TREATING INFECTIONS CAUSED BY RIFAMPICIN-RESISTANT BACTERIA 
Description Novel compounds of formulae (l)-(lll) are provided, as well as pharmaceutical compositions comprising the novel compounds. Also described are therapeutic uses of the novel compounds, in particular in relation to the treatment of infections caused by rifampicin- resistant bacteria, e.g. rifampicin-resistant tuberculosis. 
IP Reference WO2020021252 
Protection Patent granted
Year Protection Granted 2020
Licensed No
Impact NA
 
Title METHOD AND APPARATUS FOR SCANNING A SAMPLE WITH A PROBE 
Description A method of measuring a sample with a probe, the probe comprising a cantilever mount, a cantilever extending from the cantilever mount to a free end, and a probe tip carried by the free end of the cantilever, the method comprising: taking a series of sidewall measurements of a sidewall of the sample with the probe; and analysing the series of sidewall measurements to determine a characteristic of the sidewall. The sidewall measurements are taken during a sidewall measurement cycle, comprising a pair of sidewall measurement drive phases. The pair of sidewall measurement drive phases comprises a first drive phase in which the probe is driven down next to the sidewall followed by a second drive phase in which the probe is driven up next to the sidewall. During one of the sidewall measurement drive phases the probe tip interacts with the sidewall, and the series of sidewall measurements are taken by measuring an angle of the cantilever as the probe tip interacts with the sidewall during the one of the sidewall measurement drive phases. 
IP Reference WO2023057772 
Protection Patent / Patent application
Year Protection Granted 2023
Licensed Commercial In Confidence
Impact This technology is currently under development.