Sturm, A. et al. Accurate and rapid antibiotic susceptibility testing using a machine learning-assisted nanomotion technology platform. Nat. Commun. 15, 2037 (2024).
Murray, C. J. L. et al. Global burden of bacterial antimicrobial resistance in 2019: a systematic analysis. Lancet 399, 629–655 (2022).
Darby, E. M. et al. Molecular mechanisms of antibiotic resistance revisited. Nat. Rev. Microbiol. 21, 280–295 (2023).
Yau, Y. C. W. et al. Randomized controlled trial of biofilm antimicrobial susceptibility testing in cystic fibrosis patients. J. Cyst. Fibros. 14, 262–266 (2015).
Cattamanchi, A., Kyabayinze, D., Hubbard, A., Rosenthal, P. J. & Dorsey, G. Distinguishing recrudescence from reinfection in a longitudinal antimalarial drug efficacy study: comparison of results based on genotyping of msp-1, msp-2, and glurp. https://doi.org/10.4269/ajtmh.2003.68.133 (2003).
Popovici, J. et al. Recrudescence, Reinfection, or Relapse? A More Rigorous Framework to Assess Chloroquine Efficacy for Plasmodium vivax Malaria. J. Infect. Dis. 219, 315–322 (2019).
Okoro, C. K. et al. High-resolution single nucleotide polymorphism analysis distinguishes recrudescence and reinfection in recurrent invasive nontyphoidal Salmonella Typhimurium disease. Clin. Infect. Dis. 54, 955–963 (2012).
McIvor, A., Koornhof, H. & Kana, B. D. Relapse, re-infection and mixed infections in tuberculosis disease. Pathog. Dis. 75 (2017).
Xu, L. et al. Global H. pylori recurrence, recrudescence, and re-infection status after successful eradication in pediatric patients: a systematic review and meta-analysis. J. Gastroenterol. 59, 668–681 (2024).
Grant, S. S. & Hung, D. T. Persistent bacterial infections, antibiotic tolerance, and the oxidative stress response. Virulence 4, 273–283 (2013).
Ku, J. H. et al. Antibiotic resistance of urinary tract infection recurrences in a large integrated US healthcare system. J. Infect. Dis. 230, e1344–e1354 (2024).
Kadeřábková, N., Mahmood, A. J. S. & Mavridou, D. A. I. Antibiotic susceptibility testing using minimum inhibitory concentration (MIC) assays. Npj Antimicrob. Resist. 2, 1–9 (2024).
Kamaruzzaman, N. F., Kendall, S. & Good, L. Targeting the hard to reach: challenges and novel strategies in the treatment of intracellular bacterial infections. Br. J. Pharmacol. 174, 2225–2236 (2017).
Brauner, A., Fridman, O., Gefen, O. & Balaban, N. Q. Distinguishing between resistance, tolerance and persistence to antibiotic treatment. Nat. Rev. Microbiol. 14, 320–330 (2016).
Balaban, N. Q., Merrin, J., Chait, R., Kowalik, L. & Leibler, S. Bacterial Persistence as a Phenotypic Switch. Science 305, 1622–1625 (2004).
Macia, M. D., Rojo-Molinero, E. & Oliver, A. Antimicrobial susceptibility testing in biofilm-growing bacteria. Clin. Microbiol. Infect. 20, 981–990 (2014).
Aslan, H. et al. Activation of the Two-Component System LisRK Promotes Cell Adhesion and High Ampicillin Tolerance in Listeria monocytogenes. Front. Microbiol. 12, 618174 (2021).
Stupar, M. et al. TcrXY is an acid-sensing two-component transcriptional regulator of Mycobacterium tuberculosis required for persistent infection. Nat. Commun. 15, 1615 (2024).
Brown, D. R. Nitrogen Starvation Induces Persister Cell Formation in Escherichia coli. J. Bacteriol. 201, e00622–18 (2019).
Moscoso, J. A. et al. The Diguanylate Cyclase SadC Is a Central Player in Gac/Rsm-Mediated Biofilm Formation in Pseudomonas aeruginosa. J. Bacteriol. 196, 4081–4088 (2014).
Song, H., Li, Y. & Wang, Y. Two-component system GacS/GacA, a global response regulator of bacterial physiological behaviors. Eng. Microbiol. 3, 100051 (2023).
Sultan, M., Arya, R. & Kim, K. K. Roles of Two-Component Systems in Pseudomonas aeruginosa Virulence. Int. J. Mol. Sci. 22, 12152 (2021).
Ronneau, S. & Helaine, S. Clarifying the Link between Toxin–Antitoxin Modules and Bacterial Persistence. J. Mol. Biol. 431, 3462–3471 (2019).
Keren, I., Shah, D., Spoering, A., Kaldalu, N. & Lewis, K. Specialized Persister Cells and the Mechanism of Multidrug Tolerance in Escherichia coli. J. Bacteriol. 186, 8172–8180 (2004).
Tripathi, A., Dewan, P. C., Siddique, S. A. & Varadarajan, R. MazF-induced Growth Inhibition and Persister Generation in Escherichia coli. J. Biol. Chem. 289, 4191–4205 (2014).
LeRoux, M., Culviner, P. H., Liu, Y. J., Littlehale, M. L. & Laub, M. T. Stress Can Induce Transcription of Toxin-Antitoxin Systems without Activating Toxin. Mol. Cell 79, 280–292.e8 (2020).
Kawano, M., Aravind, L. & Storz, G. An antisense RNA controls synthesis of an SOS-induced toxin evolved from an antitoxin. Mol. Microbiol. 64, 738–754 (2007).
Radzikowski, J. L. et al. Bacterial persistence is an active σS stress response to metabolic flux limitation. Mol. Syst. Biol. 12, 882 (2016).
Murakami, K. et al. Role for rpoS gene of Pseudomonas aeruginosa in antibiotic tolerance. FEMS Microbiol. Lett. 242, 161–167 (2005).
Mendhe, S., Badge, A., Ugemuge, S. & Chandi, D. Impact of Biofilms on Chronic Infections and Medical Challenges. Cureus https://doi.org/10.7759/cureus.48204 (2023).
Flemming, H.-C. & Wingender, J. The biofilm matrix. Nat. Rev. Microbiol. 8, 623–633 (2010).
Liu, H. Y., Prentice, E. L. & Webber, M. A. Mechanisms of antimicrobial resistance in biofilms. Npj Antimicrob. Resist. 2, 27 (2024).
Shree, P., Singh, C. K., Sodhi, K. K., Surya, J. N. & Singh, D. K. Biofilms: Understanding the structure and contribution towards bacterial resistance in antibiotics. Med. Microecol. 16, 100084 (2023).
Yan, J. & Bassler, B. L. Surviving as a Community: Antibiotic Tolerance and Persistence in Bacterial Biofilms. Cell Host Microbe 26, 15–21 (2019).
Wivagg, C. N., Bhattacharyya, R. P. & Hung, D. T. Mechanisms of β-lactam killing and resistance in the context of Mycobacterium tuberculosis. J. Antibiot. ((Tokyo)) 67, 645–654 (2014).
Hengzhuang, W., Wu, H., Ciofu, O., Song, Z. & Høiby, N. Pharmacokinetics/Pharmacodynamics of Colistin and Imipenem on Mucoid and Nonmucoid Pseudomonas aeruginosa Biofilms. Antimicrob. Agents Chemother. 55, 4469–4474 (2011).
Bottery, M. J., Pitchford, J. W. & Friman, V.-P. Ecology and evolution of antimicrobial resistance in bacterial communities. ISME J. 15, 939–948 (2021).
Nishino, K., Yamasaki, S., Nakashima, R., Zwama, M. & Hayashi-Nishino, M. Function and Inhibitory Mechanisms of Multidrug Efflux Pumps. Front. Microbiol. 12, 737288 (2021).
Pu, Y. et al. Enhanced Efflux Activity Facilitates Drug Tolerance in Dormant Bacterial Cells. Mol. Cell 62, 284–294 (2016).
Byrd, B. A. et al. The AcrAB-TolC Efflux Pump Impacts Persistence and Resistance Development in Stationary-Phase Escherichia coli following Delafloxacin Treatment. Antimicrob. Agents Chemother. 65, e00281–21 (2021).
Martini, C. L. et al. Cellular Growth Arrest and Efflux Pumps Are Associated With Antibiotic Persisters in Streptococcus pyogenes Induced in Biofilm-Like Environments. Front. Microbiol. 12, 716628 (2021).
Anderson, G., Dodson, K., Hooton, T. & Hultgren, S. Intracellular bacterial communities of uropathogenic in urinary tract pathogenesis. Trends Microbiol. 12, 424–430 (2004).
Robino, L. et al. Presence of intracellular bacterial communities in uroepithelial cells, a potential reservoir in symptomatic and non-symptomatic people. BMC Infect. Dis. 24, 590 (2024).
Morrison, J. J. et al. Metabolic flux regulates growth transitions and antibiotic tolerance in uropathogenic Escherichia coli. J. Bacteriol. 206, e00162–24 (2024).
MacNair, C. R. & Tan, M. The role of bacterial membrane vesicles in antibiotic resistance. Ann. N. Y. Acad. Sci. 1519, 63–73 (2023).
Schwechheimer, C. & Kuehn, M. J. Outer-membrane vesicles from Gram-negative bacteria: biogenesis and functions. Nat. Rev. Microbiol. 13, 605–619 (2015).
Cao, Y. & Lin, H. Characterization and function of membrane vesicles in Gram-positive bacteria. Appl. Microbiol. Biotechnol. 105, 1795–1801 (2021).
Hua, Y. et al. Outer membrane vesicles-transmitted virulence genes mediate the emergence of new antimicrobial-resistant hypervirulent Klebsiella pneumoniae. Emerg. Microbes Infect. 11, 1281–1292 (2022).
Lehmkuhl, J. et al. Role of membrane vesicles in the transmission of vancomycin resistance in Enterococcus faecium. Sci. Rep. 14, 1895 (2024).
Rumbo, C. et al. Horizontal Transfer of the OXA-24 Carbapenemase Gene via Outer Membrane Vesicles: a New Mechanism of Dissemination of Carbapenem Resistance Genes in Acinetobacter baumannii. Antimicrob. Agents Chemother. 55, 3084–3090 (2011).
Kim, S. W. et al. The Importance of Porins and β-Lactamase in Outer Membrane Vesicles on the Hydrolysis of β-Lactam Antibiotics. Int. J. Mol. Sci. 21, 2822 (2020).
Dhital, S. et al. Neisseria gonorrhoeae -derived outer membrane vesicles package β-lactamases to promote antibiotic resistance. microLife 3, uqac013 (2022).
Olovo, C. V., Wiredu Ocansey, D. K., Ji, Y., Huang, X. & Xu, M. Bacterial membrane vesicles in the pathogenesis and treatment of inflammatory bowel disease. Gut Microbes 16, 2341670 (2024).
Murray, B. O., Dawson, R. A., Alsharaf, L. M. & Anne Winter, J. Protective effects of Helicobacter pylori membrane vesicles against stress and antimicrobial agents. Microbiology 166, 751–758 (2020).
Kulkarni, H. M., Nagaraj, R. & Jagannadham, M. V. Protective role of E. coli outer membrane vesicles against antibiotics. Microbiol. Res. 181, 1–7 (2015).
Manning, A. J. & Kuehn, M. J. Contribution of bacterial outer membrane vesicles to innate bacterial defense. BMC Microbiol. 11, 258 (2011).
Andreoni, F. et al. Antibiotics Stimulate Formation of Vesicles in Staphylococcus aureus in both Phage-Dependent and -Independent Fashions and via Different Routes. Antimicrob. Agents Chemother. 63, e01439–18 (2019).
Liu, X. et al. Research Progress on Bacterial Membrane Vesicles and Antibiotic Resistance. Int. J. Mol. Sci. 23, 11553 (2022).
Li, Q. et al. Sub-MIC Antibiotics Modulate Productions of Outer Membrane Vesicles in Tigecycline-Resistant Escherichia coli. Antibiotics 13, 276 (2024).
Mendez, J. A. et al. Extracellular Proteome of a Highly Invasive Multidrug-resistant Clinical Strain of Acinetobacter baumannii. J. Proteome Res. 11, 5678–5694 (2012).
Yonezawa, H. et al. Analysis of outer membrane vesicle protein involved in biofilm formation of Helicobacter pylori. Anaerobe 17, 388–390 (2011).
Zhao, Z. et al. Regulation of the formation and structure of biofilms by quorum sensing signal molecules packaged in outer membrane vesicles. Sci. Total Environ. 806, 151403 (2022).
Mickiewicz, K. M. et al. Possible role of L-form switching in recurrent urinary tract infection. Nat. Commun. 10, 4379 (2019).
Kilcher, S. & Loessner, M. J. Engineering Bacteriophages as Versatile Biologics. Trends Microbiol. 27, 355–367 (2019).
Kawai, Y. et al. Cell Growth of Wall-Free L-Form Bacteria Is Limited by Oxidative Damage. Curr. Biol. 25, 1613–1618 (2015).
Day, N. J., Santucci, P. & Gutierrez, M. G. Host cell environments and antibiotic efficacy in tuberculosis. Trends Microbiol. 32, 270–279 (2024).
Walsh, J. et al. Impact of host and environmental factors on β-glucuronidase enzymatic activity: implications for gastrointestinal serotonin. Am. J. Physiol. -Gastrointest. Liver Physiol. 318, G816–G826 (2020).
Nussbaumer-Pröll, A. K. et al. Impact of erythrocytes on bacterial growth and antimicrobial activity of selected antibiotics. Eur. J. Clin. Microbiol. Infect. Dis. 38, 485–495 (2019).
Ahmed, H., Bergmann, F. & Zeitlinger, M. Protein Binding in Translational Antimicrobial Development-Focus on Interspecies Differences. Antibiotics 11, 923 (2022).
Ledger, E. V. K., Mesnage, S. & Edwards, A. M. Human serum triggers antibiotic tolerance in Staphylococcus aureus. Nat. Commun. 13, 2041 (2022).
Lin, Q., Pilewski, J. M. & Di, Y. P. Acidic Microenvironment Determines Antibiotic Susceptibility and Biofilm Formation of Pseudomonas aeruginosa. Front. Microbiol. 12, 747834 (2021).
Xu, Y. et al. An acid-tolerance response system protecting exponentially growing Escherichia coli. Nat. Commun. 11, 1496 (2020).
Loffredo, M. R. et al. The pH-Insensitive Antimicrobial and Antibiofilm Activities of the Frog Skin Derived Peptide Esc(1-21): Promising Features for Novel Anti-Infective Drugs. Antibiotics 13, 701 (2024).
Martín-Gutiérrez, G. et al. Urinary Tract Physiological Conditions Promote Ciprofloxacin Resistance in Low-Level-Quinolone-Resistant Escherichia coli. Antimicrob. Agents Chemother. 60, 4252–4258 (2016).
Cunha, B. A. An infectious disease and pharmacokinetic perspective on oral antibiotic treatment of uncomplicated urinary tract infections due to multidrug-resistant Gram-negative uropathogens: the importance of urinary antibiotic concentrations and urinary pH. Eur. J. Clin. Microbiol. Infect. Dis. 35, 521–526 (2016).
Herrera-Espejo, S. et al. Acidic Urine pH and Clinical Outcome of Lower Urinary Tract Infection in Kidney Transplant Recipients Treated with Ciprofloxacin and Fosfomycin. Antibiotics 13, 116 (2024).
Kincses, A. et al. The Relationship between Antibiotic Susceptibility and pH in the Case of Uropathogenic Bacteria. Antibiotics 10, 1431 (2021).
Abbott, I. J. et al. Evaluation of pooled human urine and synthetic alternatives in a dynamic bladder infection in vitro model simulating oral fosfomycin therapy. J. Microbiol. Methods 171, 105861 (2020).
Neve, R. L., Carrillo, B. D. & Phelan, V. V. Impact of Artificial Sputum Medium Formulation on Pseudomonas aeruginosa Secondary Metabolite Production. J. Bacteriol. 203, https://doi.org/10.1128/jb.00250-21 (2021).
Tognon, M., Köhler, T., Luscher, A. & van Delden, C. Transcriptional profiling of Pseudomonas aeruginosa and Staphylococcus aureus during in vitro co-culture. BMC Genomics 20, 30 (2019).
Chapot, V., Effenberg, L., Dohmen-Ruetten, J., Buer, J. & Kehrmann, J. Evaluation of the Accelerate Pheno System for Rapid Identification and Antimicrobial Susceptibility Testing of Positive Blood Culture Bottles Inoculated with Primary Sterile Specimens from Patients with Suspected Severe Infections. J. Clin. Microbiol. 59, https://doi.org/10.1128/jcm.02637-20 (2021).
Alonso-Tarrés, C. et al. Bacteriuria and phenotypic antimicrobial susceptibility testing in 45 min by point-of-care Sysmex PA-100 System: first clinical evaluation. Eur. J. Clin. Microbiol. Infect. Dis. 43, 1533–1543 (2024).
Ersoy, S. C. et al. Correcting a fundamental flaw in the paradigm for antimicrobial susceptibility testing. EBioMedicine 20, 173–181 (2017).
Heithoff, D. M. et al. Re-evaluation of FDA-approved antibiotics with increased diagnostic accuracy for assessment of antimicrobial resistance. Cell Rep. Med. 4, 101023 (2023).
Baker, E. J., Allcott, G., Molloy, A. & Cox, J. A. G. Cystic fibrosis sputum media induces an overall loss of antibiotic susceptibility in Mycobacterium abscessus. Npj Antimicrob. Resist. 2, 1–8 (2024).
Cornforth, D. M. et al. Pseudomonas aeruginosa transcriptome during human infection. Proc. Natl. Acad. Sci. USA 115, E5125–E5134 (2018).
Cottell, J. L. & Webber, M. A. Experiences in fosfomycin susceptibility testing and resistance mechanism determination in Escherichia coli from urinary tract infections in the UK. J. Med. Microbiol. 68, 161–168 (2019).
Ciofu, O., Moser, C., Jensen, P. Ø & Høiby, N. Tolerance and resistance of microbial biofilms. Nat. Rev. Microbiol. 20, 621–635 (2022).
Padron, G. C., Shuppara, A. M., Palalay, J.-J. S., Sharma, A. & Sanfilippo, J. E. Bacteria in fluid flow. J. Bacteriol. 205, e00400–e00422 (2023).
Perry, E. K. & Tan, M.-W. Bacterial biofilms in the human body: prevalence and impacts on health and disease. Front. Cell. Infect. Microbiol. 13, 1237164 (2023).
Römling, U. & Balsalobre, C. Biofilm infections, their resilience to therapy and innovative treatment strategies. J. Intern. Med. 272, 541–561 (2012).
Sharma, D., Misba, L. & Khan, A. U. Antibiotics versus biofilm: an emerging battleground in microbial communities. Antimicrob. Resist. Infect. Control 8, 76 (2019).
Guzmán-Soto, I. et al. Mimicking biofilm formation and development: Recent progress in in vitro and in vivo biofilm models. iScience 24, 102443 (2021).
Rumbaugh, K. P. & Whiteley, M. Towards improved biofilm models. Nat. Rev. Microbiol. 23, 57–66 (2025).
Coenye, T. Biofilm antimicrobial susceptibility testing: where are we and where could we be going?. Clin. Microbiol. Rev. 36, e00024–23 (2023).
Thieme, L. et al. MBEC Versus MBIC: the Lack of Differentiation between Biofilm Reducing and Inhibitory Effects as a Current Problem in Biofilm Methodology. Biol. Proced. Online 21, 18 (2019).
Cruz, C. D., Shah, S. & Tammela, P. Defining conditions for biofilm inhibition and eradication assays for Gram-positive clinical reference strains. BMC Microbiol. 18, 173 (2018).
Bjarnsholt, T. et al. The in vivo biofilm. Trends Microbiol. 21, 466–474 (2013).
Cometta, S., Hutmacher, D. W. & Chai, L. In vitro models for studying implant-associated biofilms – A review from the perspective of bioengineering 3D microenvironments. Biomaterials 309, 122578 (2024).
Vyas, H. K. N., Xia, B. & Mai-Prochnow, A. Clinically relevant in vitro biofilm models: a need to mimic and recapitulate the host environment. Biofilm 4, 100069 (2022).
Han, Q. et al. Regrowth of microcosm biofilms on titanium surfaces after various antimicrobial treatments. Front. Microbiol. 10, 2693 (2019).
Maset, R. G. et al. Combining SNAPs with antibiotics shows enhanced synergistic efficacy against S. aureus and P. aeruginosa biofilms. Npj Biofilms Microbiomes 9, 1–17 (2023).
Jafari, N. V. & Rohn, J. L. An immunoresponsive three-dimensional urine-tolerant human urothelial model to study urinary tract infection. Front. Cell. Infect. Microbiol. 13, 1128132 (2023).
Flores, C. et al. A human urothelial microtissue model reveals shared colonization and survival strategies between uropathogens and commensals. Sci. Adv. 9, eadi9834 (2023).
Sharma, K. et al. Dynamic persistence of UPEC intracellular bacterial communities in a human bladder-chip model of urinary tract infection. eLife 10, e66481 (2021).
Murray, B. O. et al. Recurrent Urinary Tract Infection: A Mystery in Search of Better Model Systems. Front. Cell. Infect. Microbiol. 11, 691210 (2021).
Mayr, F. B., Yende, S. & Angus, D. C. Epidemiology of severe sepsis. Virulence 5, 4–11 (2014).
Foxman, B. Urinary Tract Infection Syndromes: Occurrence, Recurrence, Bacteriology, Risk Factors, and Disease Burden. Infect. Dis. Clin. North Am. 28, 1–13 (2014).
Doern, G. V. & Brecher, S. M. The Clinical Predictive Value (or Lack Thereof) of the Results of In Vitro Antimicrobial Susceptibility Tests. J. Clin. Microbiol. 49, S11–S14 (2011).
Cui, S. & Kim, E. Quorum sensing and antibiotic resistance in polymicrobial infections. Commun. Integr. Biol. 17, 2415598 (2024).
Yu, V. L. et al. An International Prospective Study of Pneumococcal Bacteremia: Correlation with In Vitro Resistance, Antibiotics Administered, and Clinical Outcome. Clin. Infect. Dis. 37, 230–237 (2003).
Somayaji, R. et al. Antimicrobial susceptibility testing (AST) and associated clinical outcomes in individuals with cystic fibrosis: A systematic review. J. Cyst. Fibros. 18, 236–243 (2019).
Bjarnsholt, T. et al. Pseudomonas aeruginosa biofilms in the respiratory tract of cystic fibrosis patients. Pediatr. Pulmonol. 44, 547–558 (2009).
Harrington, N. E., Sweeney, E. & Harrison, F. Building a better biofilm – Formation of in vivo-like biofilm structures by Pseudomonas aeruginosa in a porcine model of cystic fibrosis lung infection. Biofilm 2, 100024 (2020).
Baltimore, R. S., Christie, C. D. & Smith, G. J. Immunohistopathologic localization of Pseudomonas aeruginosa in lungs from patients with cystic fibrosis. Implications for the pathogenesis of progressive lung deterioration. Am. Rev. Respir. Dis. 140, 1650–1661 (1989).
M02 Ed14 | Performance Standards for Antimicrobial Disk Susceptibility Tests, 14th Edition. Clinical & Laboratory Standards Institute https://clsi.org/standards/products/microbiology/documents/m02/
Kowalska-Krochmal, B. & Dudek-Wicher, R. The Minimum Inhibitory Concentration of Antibiotics: Methods, Interpretation, Clinical Relevance. Pathogens 10, 165 (2021).
Balouiri, M., Sadiki, M. & Ibnsouda, S. K. Methods for in vitro evaluating antimicrobial activity: A review. J. Pharm. Anal. 6, 71 (2016).
Gajic, I. et al. Antimicrobial Susceptibility Testing: A Comprehensive Review of Currently Used Methods. Antibiotics 11, 427 (2022).
Ishak, A., Mazonakis, N., Spernovasilis, N., Akinosoglou, K. & Tsioutis, C. Bactericidal versus bacteriostatic antibacterials: clinical significance, differences and synergistic potential in clinical practice. J. Antimicrob. Chemother. 80, 1–17 (2025).
Zhang, Y., Kepiro, I., Ryadnov, M. G. & Pagliara, S. Single Cell Killing Kinetics Differentiate Phenotypic Bacterial Responses to Different Antibacterial Classes. Microbiol. Spectr. 11, e03667–22 (2023).
Levison, M. E. & Levison, J. H. Pharmacokinetics and Pharmacodynamics of Antibacterial Agents. Infect. Dis. Clin. North Am. 23, 791–vii (2009).
Bjarnsholt, T. et al. The importance of understanding the infectious microenvironment. Lancet Infect. Dis. 22, e88–e92 (2022).
Hassall, J., Coxon, C., Patel, V. C., Goldenberg, S. D. & Sergaki, C. Limitations of current techniques in clinical antimicrobial resistance diagnosis: examples and future prospects. Npj Antimicrob. Resist. 2, 16 (2024).
M26-A: Methods for Determining Bactericidal Activity of Antimicrobial Agents; Approved Guideline.
Foerster, S., Unemo, M., Hathaway, L. J., Low, N. & Althaus, C. L. Time-kill curve analysis and pharmacodynamic modelling for in vitro evaluation of antimicrobials against Neisseria gonorrhoeae. BMC Microbiol. 16, 216 (2016).
Antibiotic tolerance among clinical isolates: mechanisms, detection, prevalence, and significance | Clinical Microbiology Reviews. https://journals.asm.org/doi/10.1128/cmr.00106-24.
Mueller, M., De La Peña, A. & Derendorf, H. Issues in Pharmacokinetics and Pharmacodynamics of Anti-Infective Agents: Kill Curves versus MIC. Antimicrob. Agents Chemother. 48, 369–377 (2004).
Gumbo, T. et al. Hollow-fibre system model of tuberculosis reproducibility and performance specifications for best practice in drug and combination therapy development. J. Antimicrob. Chemother. 78, 953–964 (2023).
Hammond, R. J. H. Using Hollow Fiber to Model Treatment of Antimicrobial-Resistant Organisms. Methods Mol. Biol. Clifton NJ 2833, 57–64 (2024).
Kloprogge, F., Hammond, R., Kipper, K., Gillespie, S. H. & Della Pasqua, O. Mimicking in-vivo exposures to drug combinations in-vitro: anti-tuberculosis drugs in lung lesions and the hollow fiber model of infection. Sci. Rep. 9, 13228 (2019).
Narasimhan, V. et al. Nucleic Acid Amplification-Based Technologies (NAAT)—Toward Accessible, Autonomous, and Mobile Diagnostics. Adv. Mater. Technol. 8, 2300230 (2023).
Luo, J., Yu, J., Yang, H. & Wei, H. Parallel susceptibility testing of bacteria through culture-quantitative PCR in 96-well plates. Int. J. Infect. Dis. 70, 86–92 (2018).
Framing Bacterial Genomics for Public Health (Care) | Journal of Clinical Microbiology. https://journals.asm.org/doi/10.1128/jcm.00135-21.
Weinmaier, T. et al. Validation and Application of Long-Read Whole-Genome Sequencing for Antimicrobial Resistance Gene Detection and Antimicrobial Susceptibility Testing. Antimicrob. Agents Chemother. 67, e01072–22 (2022).
Tamae, C. et al. Determination of antibiotic hypersensitivity among 4,000 single-gene-knockout mutants of Escherichia coli. J. Bacteriol. 190, 5981–5988 (2008).
Suzuki, S., Horinouchi, T. & Furusawa, C. Prediction of antibiotic resistance by gene expression profiles. Nat. Commun. 5, 5792 (2014).
Ellington, M. J. et al. The role of whole genome sequencing in antimicrobial susceptibility testing of bacteria: report from the EUCAST Subcommittee. Clin. Microbiol. Infect. 23, 2–22 (2017).
Yoon, E.-J. & Jeong, S. H. MALDI-TOF mass spectrometry technology as a tool for the rapid diagnosis of antimicrobial resistance in bacteria. Antibiot. Basel Switz. 10, 982 (2021).
Wieser, A., Schneider, L., Jung, J. & Schubert, S. MALDI-TOF MS in microbiological diagnostics-identification of microorganisms and beyond (mini review). Appl. Microbiol. Biotechnol. 93, 965–974 (2012).
Oviaño, M. & Rodríguez-Sánchez, B. MALDI-TOF mass spectrometry in the 21st century clinical microbiology laboratory. Enfermedades Infecc. Microbiol. Clin. Engl. Ed 39, 192–200 (2021).
Idelevich, E. A. & Becker, K. Matrix-assisted laser desorption ionization-time of flight mass spectrometry for antimicrobial susceptibility testing. J. Clin. Microbiol. 59, e0181419 (2021).
Fedrigo, N. H. et al. Pharmacodynamic evaluation of fosfomycin against Escherichia coli and Klebsiella spp. from urinary tract infections and the influence of pH on fosfomycin activities. Antimicrob. Agents Chemother. 61, e02498–16 (2017).
Debets-Ossenkopp, Y. J. & MacLaren, D. M. Effect of an acidic environment on the susceptibility of helicobacterpyiori to trospectomycin and other antimicrobial agents. Eur. J. Clin. Microbiol. Infect. Dis. 14, 353–355 (1995).
Aust, A.-C. et al. Influence of kidney environment parameters on antibiotic efficacy against uropathogenic Escherichia coli. Eur. Urol. Focus 10, 742–750 (2024).
Withman, B., Gunasekera, T. S., Beesetty, P., Agans, R. & Paliy, O. Transcriptional responses of uropathogenic Escherichia coli to increased environmental osmolality caused by salt or urea. Infect. Immun. 81, 80–89 (2013).
Landry, R. M., An, D., Hupp, J. T., Singh, P. K. & Parsek, M. R. Mucin–Pseudomonas aeruginosa interactions promote biofilm formation and antibiotic resistance. Mol. Microbiol. 59, 142–151 (2006).
Vasiljevs, S., Gupta, A. & Baines, D. Effect of glucose on growth and co-culture of Staphylococcus aureus and Pseudomonas aeruginosa in artificial sputum medium. Heliyon 9, e21469 (2023).
Dörr, T., Lewis, K. & Vulić, M. SOS response induces persistence to fluoroquinolones in Escherichia coli. PLoS Genet. 5, e1000760 (2009).
Peyrusson, F. et al. Intracellular Staphylococcus aureus persisters upon antibiotic exposure. Nat. Commun. 11, 2200 (2020).
Michiels, J. E., Van Den Bergh, B., Verstraeten, N., Fauvart, M. & Michiels, J. In vitro emergence of high persistence upon periodic aminoglycoside challenge in the ESKAPE pathogens. Antimicrob. Agents Chemother. 60, 4630–4637 (2016).
Brinkman, F. S. L., Macfarlane, E. L. A., Warrener, P. & Hancock, R. E. W. Evolutionary Relationships among Virulence-Associated Histidine Kinases. Infect. Immun. 69, 5207–5211 (2001).
Murtha, A. N. et al. High-level carbapenem tolerance requires antibiotic-induced outer membrane modifications. PLoS Pathog. 18, e1010307 (2022).
Nalca, Y. et al. Quorum-sensing antagonistic activities of azithromycin in Pseudomonas aeruginosa PAO1: a global approach. Antimicrob. Agents Chemother. 50, 1680–1688 (2006).
Hentzer, M. Attenuation of Pseudomonas aeruginosa virulence by quorum sensing inhibitors. EMBO J. 22, 3803–3815 (2003).
Leung, V. & Lévesque, C. M. A stress-inducible quorum-sensing peptide mediates the formation of persister cells with noninherited multidrug tolerance. J. Bacteriol. 194, 2265–2274 (2012).
Muthuramalingam, M., White, J. C., Murphy, T., Ames, J. R. & Bourne, C. R. The toxin from a ParDE toxin-antitoxin system found in Pseudomonas aeruginosa offers protection to cells challenged with anti-gyrase antibiotics. Mol. Microbiol. 111, 441–454 (2019).
Choudhary, E., Sharma, R., Kumar, Y. & Agarwal, N. Conditional silencing by CRISPRi reveals the role of DNA gyrase in formation of drug-tolerant persister population in Mycobacterium tuberculosis. Front. Cell. Infect. Microbiol. 9, 70 (2019).
Bryson, D., Hettle, A. G., Boraston, A. B. & Hobbs, J. K. Clinical mutations that partially activate the stringent response confer multidrug tolerance in Staphylococcus aureus. Antimicrob. Agents Chemother. 64, e02103–e02119 (2020).
Dutta, N. K. et al. Inhibiting the stringent response blocks Mycobacterium tuberculosis entry into quiescence and reduces persistence. Sci. Adv. 5, eaav2104 (2019).
Anderl, J. N., Zahller, J., Roe, F. & Stewart, P. S. Role of Nutrient limitation and stationary-phase existence in Klebsiella pneumoniae biofilm resistance to ampicillin and ciprofloxacin. Antimicrob. Agents Chemother. 47, 1251–1256 (2003).
Adams, K. N. et al. Drug tolerance in replicating mycobacteria mediated by a macrophage-induced efflux mechanism. Cell 145, 39–53 (2011).
Srinivasan, V. B. & Rajamohan, G. KpnEF, a New Member of the Klebsiella pneumoniae cell envelope stress response regulon, Is an SMR-type efflux pump involved in broad-spectrum antimicrobial resistance. Antimicrob. Agents Chemother. 57, 4449–4462 (2013).
Kaiser, P. et al. Cecum lymph node dendritic cells harbor slow-growing bacteria phenotypically tolerant to antibiotic treatment. PLoS Biol. 12, e1001793 (2014).
Maredia, R. et al. Vesiculation from Pseudomonas aeruginosa under SOS. Sci. World J. 2012, 1–18 (2012).
Kim, S. W. et al. Significant increase in the secretion of extracellular vesicles and antibiotics resistance from methicillin-resistant Staphylococcus aureus induced by ampicillin stress. Sci. Rep. 10, 21066 (2020).
Zheng, Y., Cai, Y., Sun, T., Li, G. & An, T. Response mechanisms of resistance in L-form bacteria to different target antibiotics: Implications from oxidative stress to metabolism. Environ. Int. 187, 108729 (2024).
Merritt, J. H., Kadouri, D. E. & O’Toole, G. A. Growing and analyzing static biofilms. Curr. Protoc. Microbiol. 0 1, Unit-1B.1 (2005).
Crivello, G., Fracchia, L., Ciardelli, G., Boffito, M. & Mattu, C. In vitro models of bacterial biofilms: innovative tools to improve understanding and treatment of infections. Nanomaterials 13, 904 (2023).
Straub, H. et al. A microfluidic platform for in situ investigation of biofilm formation and its treatment under controlled conditions. J. Nanobiotechnol.18, 166 (2020).
Gomes, I. B. et al. Standardized reactors for the study of medical biofilms: a review of the principles and latest modifications. Crit. Rev. Biotechnol. 38, 657–670 (2018).
Lee, J.-H., Kaplan, J. B. & Lee, W. Y. Microfluidic devices for studying growth and detachment of Staphylococcus epidermidis biofilms. Biomed. Microdev.10, 489–498 (2008).
Blanco-Cabra, N. et al. A new BiofilmChip device for testing biofilm formation and antibiotic susceptibility. Npj Biofilms Microbiomes 7, 1–9 (2021).
Sriramulu, D. D., Lünsdorf, H., Lam, J. S. & Römling, U. Microcolony formation: a novel biofilm model of Pseudomonas aeruginosa for the cystic fibrosis lung. J. Med. Microbiol. 54, 667–676 (2005).
Palmer, K. L., Aye, L. M. & Whiteley, M. Nutritional cues control Pseudomonas aeruginosa multicellular behavior in cystic fibrosis sputum. J. Bacteriol. 189, 8079–8087 (2007).
Sun, Y., Dowd, S. E., Smith, E., Rhoads, D. D. & Wolcott, R. D. In vitro multispecies Lubbock chronic wound biofilm model. Wound Repair Regen. Off. Publ. Wound Heal. Soc. Eur. Tissue Repair Soc. 16, 805–813 (2008).
Werthén, M. et al. An in vitro model of bacterial infections in wounds and other soft tissues. APMIS Acta Pathol. Microbiol. Immunol. Scand. 118, 156–164 (2010).
Charles, C. A., Ricotti, C. A., Davis, S. C., Mertz, P. M. & Kirsner, R. S. Use of tissue-engineered skin to study in vitro biofilm development. Dermatol. Surg. Off. Publ. Am. Soc. Dermatol. Surg. Al 35, 1334–1341 (2009).
high-throughput microfluidic dental plaque biofilm system to visualize and quantify the effect of antimicrobials | Journal of Antimicrobial Chemotherapy | Oxford Academic. https://academic.oup.com/jac/article-abstract/68/11/2550/828884?redirectedFrom=fulltext.
Samarian, D. S., Jakubovics, N. S., Luo, T. L. & Rickard, A. H. Use of a high-throughput in vitro microfluidic system to develop oral multi-species biofilms. J. Vis. Exp. JoVE 52467 https://doi.org/10.3791/52467 (2014).
Lamret, F. et al. Human Osteoblast-Conditioned Media Can Influence Staphylococcus aureus Biofilm Formation. Int. J. Mol. Sci. 23, 14393 (2022).
Chutipongtanate, S. & Thongboonkerd, V. Systematic comparisons of artificial urine formulas for in vitro cellular study. Anal. Biochem. 402, 110–112 (2010).
Ipe, D. S. & Ulett, G. C. Evaluation of the in vitro growth of urinary tract infection-causing gram-negative and gram-positive bacteria in a proposed synthetic human urine (SHU) medium. J. Microbiol. Methods 127, 164–171 (2016).
Rimbi, P. T. et al. Enhancing a multi-purpose artificial urine for culture and gene expression studies of uropathogenic Escherichia coli strains. J. Appl. Microbiol. 135, lxae067 (2024).
Townsend, E. M., Moat, J. & Jameson, E. CAUTI’s next top model – Model dependent Klebsiella biofilm inhibition by bacteriophages and antimicrobials. Biofilm 2, 100038 (2020).
Zaborskyte, G., Wistrand-Yuen, E., Hjort, K., Andersson, D. I. & Sandegren, L. Modular 3D-printed peg biofilm device for flexible setup of surface-related biofilm studies. Front. Cell. Infect. Microbiol. 11, 802303 (2022).
Alves, D. R. et al. Development of a High-Throughput ex-Vivo Burn Wound Model Using Porcine Skin, and Its Application to Evaluate New Approaches to Control Wound Infection. Front. Cell. Infect. Microbiol. 8, 196 (2018).
Maset, R. G. et al. Combining SNAPs with antibiotics shows enhanced synergistic efficacy against S. aureus and P. aeruginosa biofilms. NPJ Biofilms Microbiomes 9, 36 (2023).
Wurbs, A. et al. A human ex vivo skin model breaking boundaries. Sci. Rep. 14, 24054 (2024).
Du, Q. et al. Candida albicans promotes tooth decay by inducing oral microbial dysbiosis. ISME J. 15, 894–908 (2021).
de Poel, E. et al. FDA-approved drug screening in patient-derived organoids demonstrates potential of drug repurposing for rare cystic fibrosis genotypes. J. Cyst. Fibros. Off. J. Eur. Cyst. Fibros. Soc. 22, 548–559 (2023).
Sollier, J. et al. Revitalizing antibiotic discovery and development through in vitro modelling of in-patient conditions. Nat. Microbiol. 9, 1–3 (2024).
Aguilar, C. et al. Organoids as host models for infection biology – a review of methods. Exp. Mol. Med. 53, 1471–1482 (2021).
Swart, A. L. et al. Pseudomonas aeruginosa breaches respiratory epithelia through goblet cell invasion in a microtissue model. Nat. Microbiol. 9, 1725–1737 (2024).
Meirelles, L. A. et al. Pseudomonas aeruginosa faces a fitness trade-off between mucosal colonization and antibiotic tolerance during airway infection. Nat. Microbiol. 9, 3284–3303 (2024).
Han, X. et al. Creating a more perfect union: modeling intestinal bacteria-epithelial interactions using organoids. Cell. Mol. Gastroenterol. Hepatol. 12, 769–782 (2021).
Anonye, B. O. et al. Probing Clostridium difficile Infection in Complex Human Gut Cellular Models. Front. Microbiol. 10, 879 (2019).
Dash, S. K., Marques, C. N. H. & Mahler, G. J. Small intestine on a chip demonstrates physiologic mucus secretion in the presence of Lacticaseibacillus rhamnosus biofilm. Biotechnol. Bioeng. 1–12 (2025).
Łaniewski, P. & Herbst-Kralovetz, M. M. Bacterial vaginosis and health-associated bacteria modulate the immunometabolic landscape in 3D model of human cervix. Npj Biofilms Microbiomes 7, 1–17 (2021).
Redman, W. K. et al. Efficacy and safety of biofilm dispersal by glycoside hydrolases in wounds. Biofilm 3, 100061 (2021).
Nissanka, M. C., Dilhari, A., Wijesinghe, G. K. & Weerasekera, M. M. Advances in experimental bladder models: bridging the gap between in vitro and in vivo approaches for investigating urinary tract infections. BMC Urol. 24, 206 (2024).
Conover, M. S., Flores-Mireles, A. L., Hibbing, M. E., Dodson, K. & Hultgren, S. J. Establishment and Characterization of UTI and CAUTI in a Mouse Model. J. Vis. Exp. JoVE e52892, https://doi.org/10.3791/52892 (2015).